CN113692460B

CN113692460B - Recombinant structural protein multifilament and its production method

Info

Publication number: CN113692460B
Application number: CN202080012298.1A
Authority: CN
Inventors: 安部佑之介; 石井秀人; 佐藤僚子; 佐藤皓斗; 安藤广忠; 狩野博司; 小鹰浩一; 小林淳
Original assignee: Spiber Inc
Current assignee: Sibabo Intellectual Property Management Contract Co
Priority date: 2019-02-07
Filing date: 2020-02-07
Publication date: 2024-04-26
Anticipated expiration: 2040-02-07
Also published as: MY209893A; JPWO2020162626A1; JP7685739B2; EP3922764A4; WO2020162626A1; US20220010460A1; EP3922764A1; CN113692460A

Abstract

The present invention aims to provide a method for producing modified fibroin multifilament with excellent productivity and the multifilament produced thereby. The multifilament of the present invention contains modified fibroin and has 100 or more constituent filaments, the coefficient of variation of elastic modulus is 15% or less, the coefficient of variation of strength is 15% or less, and the coefficient of variation of elongation is less than 33%.

Description

Recombinant structural protein multifilament yarn and method for producing the same

Technical Field

The present invention relates to a recombinant structural protein multifilament yarn and a method for producing the same.

Background

Conventionally, as artificial silk fibroin fibers, spider silk fibroin fibers, and the like, which are regenerated silk fibers, have been known, and many of these spinning methods have been reported.

For example, there is reported a multifilament yarn comprising spider silk fibroin derived from a natural spider silk fibroin structure (patent document 1), which is produced using a spinning nozzle having a number of 50 holes and is composed of 50 monofilaments.

However, there has not been reported a recombinant structural protein multifilament yarn which is mass-produced using a spinning nozzle having a larger number of holes and which comprises a plurality of monofilaments having the same number of holes as the spinning nozzle.

Prior art literature

Patent literature

Patent document 1: international publication No. 2018/053204

Disclosure of Invention

Problems to be solved by the invention

The present invention aims to provide a multifilament yarn which is composed of a plurality of monofilaments and contains a plurality of recombinant structural proteins and does not exist in the world.

Means for solving the problems

That is, the present invention relates to, for example, the following inventions.

[1] A multifilament yarn comprising modified fibroin and having 100 or more constituent filaments, a coefficient of variation of elastic modulus of 15% or less, a coefficient of variation of strength of 15% or less, and a coefficient of variation of elongation of less than 33%.

[2] The multifilament according to [1], wherein the multifilament has a coefficient of variation of elongation of 20% or less.

[3] The multifilament according to [1] or [2], wherein the coefficient of variation in elongation of the multifilament is 0.01% or more and 20% or less.

[4] The multifilament yarn according to any one of [1] to [3], wherein the elongation coefficient of the multifilament yarn has a coefficient of variation of 0.01% or more and less than 0.1%, more than 0.1% and less than 1%, more than 1% and less than 5%, more than 5% and less than 10%, more than 10% and less than 15% or more than 15% and less than 20%.

[5] The multifilament yarn according to any one of [1] to [4], wherein the multifilament yarn has a coefficient of variation in elongation of 15% or less.

[6] The multifilament yarn according to any one of [1] to [4], wherein the multifilament yarn has a coefficient of variation in elongation of 10% or less.

[7] The multifilament yarn according to any one of [1] to [6], wherein the coefficient of variation of fineness of the multifilament yarn is 20% or less.

[8] The multifilament yarn according to any one of [1] to [7], wherein the coefficient of variation in the strength of the multifilament yarn is 10% or less, and the coefficient of variation in the elastic modulus of the multifilament yarn is 10% or less.

[9] The multifilament yarn according to any one of [1] to [8], wherein the modified fibroin has an average hydrophobicity index of more than-0.8.

[10] The multifilament yarn according to any one of [1] to [9], wherein the modified silk fibroin is a modified spider silk fibroin.

[11] The multifilament yarn according to any one of [1] to [10], wherein it has a shrinkage history in which irreversible shrinkage occurs after spinning.

[12] The multifilament yarn according to [11], wherein the shrinkage history is a shrinkage history by bringing the multifilament yarn into contact with water to cause irreversible shrinkage or a shrinkage history by heat-relaxing the multifilament yarn to cause irreversible shrinkage.

[13] The multifilament according to [11] or [12], wherein the shrinkage of the multifilament having a shrinkage history of irreversible shrinkage after spinning, defined by the following formula, is 5% or less.

Shrinkage [% ] = (1- (length of multifilament when dried from wet state/length of multifilament when wet state)) ×100

[14] The multifilament yarn according to any one of [1] to [13], wherein the coefficient of variation of fineness of the multifilament yarn is 0.01 to 6.5%.

[15] The multifilament yarn according to any one of [1] to [14], wherein the strength of the multifilament yarn has a coefficient of variation of 0.01 to 3.8%.

[16] A multifilament yarn comprising a recombinant structural protein, having 100 or more constituent filaments, and having an elongation coefficient of variation of less than 33%.

[17] The multifilament yarn according to [16], wherein the recombinant structural protein satisfies the following (1) or (2).

(1) The number of amino acid residues is more than 150, the content of alanine residues is 12-40%, and the content of glycine residues is 11-55%.

(2) The total of the amino acid residue content, alanine residue content, and glycine residue content of at least 1 selected from the group consisting of serine, threonine, and tyrosine is 56% or more.

[18] The multifilament yarn according to [16] or [17], wherein the recombinant structural protein satisfies both of the above (1) and (2).

[19] The multifilament yarn according to any one of [16] to [18], wherein the recombinant structural protein has a plurality of repeating sequence units, and the number of amino acid residues of the repeating sequence units is 6 to 200.

[20] The multifilament yarn according to any one of [16] to [19], wherein the recombinant structural protein is at least 1 selected from the group consisting of fibroin, collagen, resilin, elastin and keratin, and proteins derived therefrom.

[21] The multifilament yarn according to any one of [16] to [20], wherein the recombinant structural protein comprises the motif (A) _n.

[22] A method for producing a multifilament, comprising a step of ejecting a spinning solution containing a recombinant structural protein and a solvent from a spinning nozzle having a number of holes of 100 or more to bring the spinning solution into contact with a coagulation liquid, and coagulating the recombinant structural protein.

[23] The method for producing a multifilament yarn according to [22], wherein the recombinant structural protein satisfies the following (1) or (2).

[24] The method for producing a multifilament yarn according to [22] or [23], wherein the recombinant structural protein satisfies both of the above (1) and (2).

[25] The method for producing a multifilament yarn according to any one of [22] to [24], wherein the recombinant structural protein has a plurality of repeating sequence units and the number of amino acid residues of the repeating sequence units is 6 to 200.

[26] The production method according to any one of [22] to [25], wherein the recombinant structural protein is at least 1 selected from the group consisting of fibroin, collagen, resilin, elastin and keratin, and proteins derived therefrom.

[27] The multifilament yarn according to any one of [22] to [26], wherein the recombinant structural protein comprises the motif (a) _n.

[28] A method for producing a multifilament, comprising a step of ejecting a spinning solution containing modified fibroin and a solvent from a spinning nozzle having a number of holes of 100 or more to bring the spinning solution into contact with a coagulation liquid, and coagulating the modified fibroin.

[29] The production method according to [28], wherein the number of holes of the spinning nozzle is 100 to 9000 inclusive.

[30] The production method according to any one of [28] and [29], wherein the modified fibroin has an average hydrophobicity index of more than-0.8.

[31] The method of producing according to any one of [28] to [30], wherein the modified silk fibroin is a modified spider silk fibroin.

[32] The production method according to any one of [28] to [31], wherein the above-mentioned solvent is at least 1 organic solvent selected from the group consisting of formic acid, DMSO and HFIP.

[33] The production method according to any one of [28] to [32], wherein the solidification liquid contains at least 1 component selected from the group consisting of a lower alcohol having 1 to 5 carbon atoms, a ketone, water, and an aqueous solution having a pH of 0.25 or more and a pH of 10.00 or less.

[34] The production method according to [33], wherein the content of the above-mentioned component is 70% by mass or more based on 100% by mass of the total amount of the solidification liquid.

[35] The production method according to any one of [28] to [34], wherein the solidification liquid contains at least 1 kind of a group consisting of methanol, acetone, water, an aqueous sodium chloride solution, an aqueous sodium sulfate solution, and an aqueous formic acid solution.

[36] The production method according to any one of [28] to [35], wherein the solidification liquid contains at least 1 kind of a group consisting of methanol, an aqueous sodium sulfate solution, and an aqueous formic acid solution.

[37] The method according to any one of [28] to [36], wherein the solidification liquid contains an organic solvent, and the content of the organic solvent is 30 mass% or less based on 100 mass% of the total solidification liquid.

Effects of the invention

According to the present invention, there can be provided a multifilament yarn comprising a plurality of filaments and containing a recombinant protein, which is not present in the world, and a method for producing the same. The multifilament yarn comprising a recombinant protein according to the present invention has a smaller coefficient of variation in elongation, modulus of elasticity, strength and/or fineness, and in particular, has a smaller coefficient of variation in elongation than conventional multifilament yarns (for example, patent document 1). That is, the multifilament yarn comprising recombinant protein according to the present invention has small relative variation in physical properties and extremely excellent quality stability.

Drawings

FIG. 1 is a schematic diagram showing an example of a domain sequence of an altered fibroin.

FIG. 2 is a graph showing the distribution of z/w (%) values of a naturally-derived fibroin.

FIG. 3 is a graph showing the distribution of values of x/y (%) of a naturally-derived fibroin.

FIG. 4 is a schematic diagram showing an example of the domain sequence of the modified fibroin.

FIG. 5 is a schematic diagram showing an example of the domain sequence of the modified fibroin.

Fig. 6 is an explanatory view schematically showing an example of a spinning apparatus for producing a modified fibroin multifilament yarn.

Fig. 7 is an explanatory view schematically showing an example of a spinning apparatus for producing a modified fibroin multifilament yarn.

Fig. 8 is a diagram showing an example of a change in the length of a filament caused by contact with water.

Fig. 9 is an explanatory view schematically showing an example of a production apparatus for producing the modified fibroin multifilament yarn.

Fig. 10 is an explanatory view schematically showing an example of a production apparatus for producing the modified fibroin multifilament yarn.

Fig. 11 is an explanatory view schematically showing an example of a production apparatus for producing the modified fibroin multifilament yarn.

Fig. 12 is an explanatory view showing a speed adjusting unit and a temperature adjusting unit that can be provided in the high-temperature heating furnace of fig. 11.

FIG. 13 is a photograph showing a cross section of a modified fibroin multifilament (the number of constituent filaments of the multifilament: 1,000) produced using a spinning nozzle having a hole number of 1,000.

FIG. 14 is a photograph showing a cross section of a modified fibroin multifilament (the number of constituting filaments of the multifilament: 3,000) produced using a spinning nozzle having a hole number of 3,000.

Fig. 15 is a graph showing an example of the results of the hygroscopic heat generation performance test.

Detailed Description

(Recombinant structural proteins)

The structural protein means a protein forming an active structure or a protein derived from the same. The recombinant structural protein is a structural protein produced by a genetic recombination technique. The recombinant structural protein may have an amino acid sequence of a structural protein of natural origin, or may be a modified structural protein in which a part of the amino acid sequence is modified according to the amino acid sequence of a structural protein of natural origin.

The recombinant structural protein according to the present embodiment may be a protein satisfying any one of the following (1) or (2).

In the present specification, "alanine residue content" refers to a value represented by the following formula.

Alanine residue content= (number of alanine residues contained in recombinant structural protein/number of all amino acid residues of polypeptide) ×100 (%)

In addition, the glycine residue content, serine residue content, threonine residue content and tyrosine residue content are the same as in the above formula in which alanine residues are replaced with glycine residues, serine residues, threonine residues and tyrosine residues, respectively.

The recombinant structural protein satisfying (1) may have at least 150 amino acid residues. The number of amino acid residues may be, for example, 200 or more or 250 or more, preferably 300 or more, 350 or more, 400 or more, 450 or more, or 500 or more.

The recombinant structural protein satisfying (1) is only required to have an alanine residue content of 12 to 40%. The content of the alanine residue may be, for example, 15 to 40%, 18 to 40%, 20 to 40%, or 22 to 40%.

The recombinant structural protein satisfying (1) is only required to have a glycine residue content of 11 to 55%. The glycine residue content may be, for example, 11% to 55%, 13% to 55%, 15% to 55%, 18% to 55%, 20% to 55%, 22% to 55%, or 25% to 55%.

The recombinant structural protein satisfying (2) may be one in which at least 1 amino acid residue content (i.e., serine residue content, threonine residue content, tyrosine residue content, total of serine residue content and threonine residue content, total of serine residue content and tyrosine residue content, total of threonine residue content and tyrosine residue content, total of serine residue content, threonine residue content and tyrosine residue content) or total content (total content) of alanine residue content and glycine residue content is 56% or more. The total content may be, for example, 57% or more, 58% or more, 59% or more, or 60% or more. The upper limit of the total content is not particularly limited, and may be, for example, 90% or less, 85% or less, or 80% or less.

In one embodiment, the total of serine residue content, threonine residue content, and tyrosine residue content of the recombinant structural protein satisfying (2) may be 4% or more, may be 4.5% or more, may be 5% or more, may be 5.5% or more, may be 6% or more, may be 6.5% or more, and may be 7% or more. The total of the serine residue content, threonine residue content, and tyrosine residue content may be, for example, 35% or less, 33% or less, 30% or less, 25% or less, or 20% or less.

The recombinant structural protein according to the present embodiment is preferably a protein satisfying both of the above (1) and (2). Thus, the effects of the present invention are more remarkably exhibited.

The distribution of serine residues, threonine residues, or tyrosine residues in the recombinant structural proteins according to this embodiment is average, and the total content of serine residues, threonine residues, and tyrosine residues may be 5% or more, 10% or more, or 15% or more, or 50% or less, 40% or less, 30% or less, or 20% or less in any of the consecutive 20 amino acid residues.

The recombinant structural protein according to one embodiment may be a protein having a repetitive sequence. That is, the recombinant structural protein according to the present embodiment may have a plurality of amino acid sequences (repeating sequence units) having high sequence identity in the recombinant structural protein. The amino acid sequence of the repeating sequence unit is not particularly limited, and the whole recombinant structural protein may be a protein satisfying the above (1) or (2). The number of amino acid residues in the repeating sequence unit is preferably 6 to 200. The sequence identity between the repeating sequence units may be 85% or more, 90% or more, 95% or more, 96% or more, 97% or more, 98% or more, or 99% or more, for example.

The recombinant structural protein according to one embodiment may be a protein comprising the motif of (a) _n. In the present specification, (A) _n motif means an amino acid sequence based on an alanine residue. (A) The _n motif may have an amino acid residue number of 2 to 27, or may be an integer of 2 to 20, 2 to 16, or 2 to 12. The ratio of the number of alanine residues in the motif (a) _n to the total number of amino acid residues may be 60% or more, 70% or more, 80% or more, 83% or more, 85% or more, 86% or more, 90% or more, 95% or more, or 100% (meaning that it is composed of only alanine residues).

In one embodiment, the (a) _n motif may be contained in a repeat unit. (A) The _n motif comprises mainly alanine residues, thus an alpha-helical structure or a beta-sheet structure is readily available. Since the recombinant structural protein according to the present embodiment repeatedly has these secondary structures by including the (a) _n motif in the repeat sequence unit, it is expected that the recombinant structural protein will exert high strength by these secondary structures when the recombinant structural protein is in the form of a fiber.

Examples of the recombinant structural protein include any structural protein that is preferably produced on an industrial scale, and specifically, structural proteins that can be used in industry, structural proteins that can be used in medical treatment, and the like. Specific examples of structural proteins usable in industry or medical treatment include fibroin, collagen, elastin, keratin, and proteins derived from these, which are poorly soluble proteins, and water-soluble proteins. The silk fibroin may be, for example, 1 or more selected from the group consisting of silk fibroin (silk protein), spider silk fibroin (spider silk protein), and bee silk fibroin.

The fibroin of the present embodiment includes fibroin of natural origin and modified fibroin. In the present specification, "native-derived fibroin" means fibroin having the same amino acid sequence as native-derived fibroin, and "modified fibroin" means fibroin having an amino acid sequence different from that of native-derived fibroin. In the present specification, "modified fibroin" refers to an artificially produced fibroin (artificial fibroin). The modified fibroin may be a fibroin having a domain sequence different from that of a fibroin derived from a natural source, or a fibroin having the same amino acid sequence as that of a fibroin derived from a natural source.

The fibroin according to the present embodiment is preferably spider silk fibroin (spider silk protein). Spider silk fibroin comprises natural spider silk fibroin and modified spider silk fibroin derived from natural spider silk fibroin. As the natural spider silk fibroin, for example, spider silk proteins produced by spider are cited.

The fibroin according to the present embodiment may be, for example, a fibroin comprising formula 1: [ (a) _n motif-REP ] _m or formula 2: a protein having a domain sequence represented by the [ (A) _n motif-REP ] _m-(A)_n motif. The fibroin of the present embodiment may further have an amino acid sequence (an N-terminal sequence and a C-terminal sequence) added to either one or both of the N-terminal side and the C-terminal side of the domain sequence. The N-terminal sequence and the C-terminal sequence are not limited to these, and are usually regions having no repetition of an amino acid motif characteristic in fibroin, and are composed of about 100 residues of amino acids.

Modified fibroin the modified fibroin as a raw material is not particularly limited, and may be produced by a gene recombination technique using a microorganism or the like, or may be produced synthetically. However, the modified fibroin does not include fibroin of natural origin. In addition, in the present embodiment, when the modified silk fibroin is modified spider silk fibroin, the heat insulating performance, the hygroscopic and exothermic performance, and/or the flame retardancy can be made more excellent.

The modified fibroin according to the present embodiment is represented by the following formula 1: [ (a) _n motif-REP ] _m or formula 2: a protein having a domain sequence represented by the [ (A) _n motif-REP ] _m-(A)_n motif. The modified fibroin may further have an amino acid sequence (an N-terminal sequence and a C-terminal sequence) attached to either one or both of the N-terminal side and the C-terminal side of the domain sequence. The N-terminal sequence and the C-terminal sequence are not limited to these, and are usually regions having no repetition of an amino acid motif characteristic in fibroin, and are composed of about 100 residues of amino acids.

In the present specification, "modified fibroin" refers to an artificially produced fibroin (artificial fibroin). The modified fibroin may be a fibroin having a domain sequence different from that of a fibroin derived from a natural source, or a fibroin having the same amino acid sequence as that of a fibroin derived from a natural source. In the present specification, "fibroin of natural origin" is also a protein comprising formula 1: [ (a) _n motif-REP ] _m or formula 2: a protein having a domain sequence represented by the [ (A) _n motif-REP ] _m-(A)_n motif.

"Modified fibroin" may be a fibroin that directly uses the amino acid sequence of a naturally-derived fibroin, a fibroin that is modified according to the amino acid sequence of a naturally-derived fibroin (for example, a fibroin obtained by modifying the amino acid sequence of a cloned naturally-derived fibroin) or a fibroin that is designed and synthesized artificially independent of a naturally-derived fibroin (for example, a fibroin that has a desired amino acid sequence by chemically synthesizing a nucleic acid encoding the designed amino acid sequence).

In the present specification, the "domain sequence" refers to an amino acid sequence that is a crystalline region (typically, equivalent to the (a) _n motif of an amino acid sequence) and an amorphous region (typically, equivalent to the REP of an amino acid sequence) unique to fibroin, and refers to formula 1: [ (a) _n motif-REP ] _m or formula 2: the amino acid sequence represented by the [ (A) _n motif-REP ] _m-(A)_n motif. Here, (A) _n represents an amino acid sequence mainly comprising alanine residues, and the number of amino acid residues is 2 to 27. (A) The number of amino acid residues of the _n motif may be an integer from 2 to 20, 4 to 27, 4 to 20, 8 to 20, 10 to 20, 4 to 16, 8 to 16 or 10 to 16. The ratio of the number of alanine residues in the motif (a) _n to the total number of amino acid residues may be 60% or more, 70% or more, 80% or more, 83% or more, 85% or more, 86% or more, 90% or more, 95% or more, or 100% (meaning that it is composed of only alanine residues). At least 7 of the plurality of (a) _n motifs present in the domain sequence may consist of only alanine residues. REP represents an amino acid sequence consisting of 2 to 200 amino acid residues. REP may be an amino acid sequence of 10 to 200 amino acid residues, or an amino acid sequence of 10 to 40, 10 to 60, 10 to 80, 10 to 100, 10 to 120, 10 to 140, 10 to 160 or 10 to 180 amino acid residues. m represents an integer of 2 to 300, and may be an integer of 8 to 300, 10 to 300, 20 to 300, 40 to 300, 60 to 300, 80 to 300, 10 to 200, 20 to 180, 20 to 160, 20 to 140, or 20 to 120. The two or more motifs (A) _n may have the same amino acid sequence or may have different amino acid sequences. The REPs may have the same amino acid sequence as each other or may have different amino acid sequences from each other.

The modified fibroin according to the present embodiment can be obtained, for example, by modifying a cloned gene sequence of a naturally-derived fibroin, for example, by modifying an amino acid sequence corresponding to substitution, deletion, insertion, and/or addition of one or more amino acid residues. Substitution, deletion, insertion and/or addition of amino acid residues can be performed by methods well known to those skilled in the art, such as site-directed mutagenesis. Specifically, the method can be performed according to the methods described in the documents such as Nucleic Acid res.10, 6487 (1982), methods in Enzymology,100, 448 (1983), and the like.

The fibroin of natural origin is of the formula 1: [ (a) _n motif-REP ] _m, or formula 2: the protein having the domain sequence represented by the [ (A) _n motif-REP ] _m-(A)_n motif, specifically, for example, silk fibroin produced by insects or arachnids can be exemplified.

Examples of the silk fibroin produced by insects include silk proteins produced by silkworms such as Bombyx mori (Bombyx mori), wild silkworm (Bombyx mandarina), wild silkworm (ANTHERAEA YAMAMAI), tussah (ANTERAEA PERNYI), maple silkworm (Eriogyna pyretorum), castor silkworm (Pilosamia CYNTHIA RICINI), ailanthus altissima silkworm (SAMIA CYNTHIA), camphor silkworm (Caligura japonica), indian tussah silkworm (ANTHERAEA MYLITTA), and amber silkworm (ANTHERAEA ASSAMA), and bee silk proteins produced by larvae of queen bee (VESPA SIMILLIMA xanthoptera).

More specific examples of the fibroin produced by insects include, for example, silkworm fibroin L chain (GenBank accession No. M76430 (base sequence) and AAA27840.1 (amino acid sequence)).

As a silk fibroin produced by the arachnids, examples thereof include spiders belonging to the genus Aranea (genus Araneus), such as Aralia elata, and Aralia elata (Araneus nojimai), spiders belonging to the genus New Aralia (genus Neoscona), such as New Aralia elata, bush, and New Aralia elata, and spiders belonging to the genus Agkistrodon (genus Pronus), such as cape spider, and the like spiders belonging to genus Abdominal (CYRTARACHNE), such as Bufo siccus Abdominal spider and Symphytum, spidroa kurzonera and Petasites (GASTERACANTHA), spiders belonging to genus Abdominal (Ordgarius), such as Hematopsis schwanensis and Hematopsis schwanensis, spiders belonging to genus Abdominal (Argiope), such as Octopus, and Petasites, spiders belonging to genus Abdominal (Argiope), such as Octopus, and Petasites, and the like spider belonging to genus Larix (Arachnura), spider belonging to genus Larix, brown spider She Zhu, etc., to genus Larix (Acusilas), spider belonging to genus Larix (Cytophora), such as red-cloud spider, flower-cloud spider and panchromatic cloud spider, spider belonging to genus Larix (Poltys), such as Philippine spider, octalop Ai Zhu, tetranychus, round abdomen Ai Zhu and Heidesia Ai Zhu, spider belonging to genus Larix (Cyclosa), spider silk protein produced by spider belonging to genus Larix (Chorizopes), such as Japanese Zhuang head spider, and spider belonging to genus Agrocarpus (Leucauge), such as front tooth spider, console, direct-stretch spider and scaly spider, etc, spiders belonging to genus Nephila such as Nephila and Nephia, spiders belonging to genus Maitake (Menosira) such as Nephia, spiders belonging to genus Nephia such as Nephia, spiders belonging to genus Serratia such as Arecaria, spiders belonging to genus Serratia (Dyschiriognatha), spiders belonging to genus Myrotheca such as Tacronathus, latrodes such as Tacronathus Kou Zhu, ha Shikou, and Methanosatus Kou Zhu, spiders belonging to genus Latrodes, and spiders belonging to family Celasthenops such as Araceae (TETRAGNATHIDAE). Examples of the spider silk proteins include dragline silk proteins such as MaSp (MaSp 1 and MaSp 2) and ADF (ADF 3 and ADF 4), miSp (MiSp 1 and MiSp 2), and the like.

More specific examples of spider silk proteins produced by spider silk include-3 (-3) [ from ] (GenBank accession AAC (amino acid sequence), U (base sequence)), -4 (-4) [ from ] (GenBank accession AAC47011 (amino acid sequence), U (base sequence)), 1[ from ] (GenBank accession AAC04504 (amino acid sequence), U (base sequence)), 1[ from ] (GenBank accession ABR (amino acid sequence), EF (base sequence)), 2[ from ] (GenBank accession AAL (amino acid sequence), AF (base sequence)), 1[ from ] (GenBank accession CAJ00428 (amino acid sequence), AJ (base sequence)), and 2[ (GenBank accession CAM (amino acid sequence), AM (base sequence)) ], ] (GenBank accession AAC (amino acid sequence)), and 2[ (GenBank accession AAC (amino acid sequence)), and the like.

As a more specific example of the fibroin of natural origin, fibroin in which sequence information is registered in NCBI GenBank can be further exemplified. For example, it can be confirmed by extracting a sequence in which spider silk proteins, pot glands, silk fibroin, "silk and polypeptide" or "silk and protein" are described in the DEFINITION (DEFINITION) from a sequence in which INV is included as a classification code (DIVISION) in sequence information registered in NCBI GenBank, extracting a character string of a specified product from CDS, extracting a sequence in which a specified character string is described in the TISSUE TYPE (tisset TYPE) from SOURCE (SOURCE).

The modified fibroin according to the present embodiment may be modified silk fibroin (modified silk fibroin having an amino acid sequence of fibroin produced by silkworm) or modified spider silk fibroin (modified silk fibroin having an amino acid sequence of spider silk protein produced by spider). As the modified silk fibroin, modified spider silk fibroin is preferable.

Specific examples of modified fibroin include modified spider silk fibroin derived from large silk tube traction silk protein produced by large pot glands of spiders (modified silk fibroin 1 st), modified spider silk fibroin having a domain sequence with a reduced glycine residue content (modified silk fibroin 2 nd), modified spider silk fibroin having a domain sequence with a reduced (a) _n motif content (modified silk fibroin 3 rd), modified spider silk fibroin having a reduced glycine residue content and (a) _n motif content (modified silk fibroin 4 th), modified spider silk fibroin having a domain sequence with a region with a large hydrophobicity index locally (modified silk fibroin 5 th), and modified spider silk fibroin having a domain sequence with a reduced glutamine residue content (modified silk fibroin 6 th). These modified fibroin are excellent in flame retardancy, hygroscopic and exothermic properties, and thermal insulation properties, and are also suitable for use in fireproof clothing (for example, firefighters and rescue clothing), fireproof gloves (for example, for laboratory use, industrial use, and cooking use), gloves, scarves, sweaters, outerwear, jackets, and other cold-proof clothing (cold-proof clothing materials), cotton wool for cold-proof clothing materials, underwear, sportswear, shirts, bedding, and cotton wool for bedding.

The modified fibroin of item 1 may include the modified fibroin of formula 1: protein of domain sequence shown in [ (A) _n motif-REP ] _m. In the modified fibroin of item 1, the amino acid residue number n of the motif (a) _n is preferably an integer of 3 to 20, more preferably an integer of 4 to 20, still more preferably an integer of 8 to 20, still more preferably an integer of 10 to 20, still more preferably an integer of 4 to 16, particularly preferably an integer of 8 to 16, and most preferably an integer of 10 to 16. In the modified fibroin of the 1 st, the number of amino acid residues constituting REP in the formula 1 is preferably 10 to 200 residues, more preferably 10 to 150 residues, further preferably 20 to 100 residues, still further preferably 20 to 75 residues. In the 1 st modified fibroin, formula 1: the total number of glycine residues, serine residues and alanine residues contained in the amino acid sequence represented by [ (A) _n motif-REP ] _m is preferably 40% or more, more preferably 60% or more, still more preferably 70% or more, based on the total number of amino acid residues.

The 1 st modified fibroin can be the following polypeptide: comprising formula 1: the unit of the amino acid sequence shown in [ (A) _n motif-REP ] _m ], and the C-terminal sequence is the amino acid sequence shown in any one of SEQ ID Nos. 1 to 3 or the amino acid sequence having homology of 90% or more with the amino acid sequence shown in any one of SEQ ID Nos. 1 to 3.

The amino acid sequence shown in SEQ ID NO. 1 is identical to an amino acid sequence consisting of 50 residues of the C-terminal amino acid of the amino acid sequence of ADF3 (GI: 1263287, NCBI), the amino acid sequence shown in SEQ ID NO. 2 is identical to an amino acid sequence obtained by removing 20 residues from the C-terminal amino acid sequence shown in SEQ ID NO. 1, and the amino acid sequence shown in SEQ ID NO.3 is identical to an amino acid sequence obtained by removing 29 residues from the C-terminal amino acid sequence shown in SEQ ID NO. 1.

More specific examples of the modified fibroin of item 1 include modified fibroin having an amino acid sequence having 90% or more sequence identity with the amino acid sequence shown in (1-i) SEQ ID NO. 4 (recombiant SPIDER SILK protein ADF3KAILARGENRSH 1) or the amino acid sequence shown in (1-ii) SEQ ID NO. 4. The sequence identity is preferably 95% or more.

The amino acid sequence shown in SEQ ID NO. 4 was obtained by mutating the amino acid sequence of ADF3, to which an amino acid sequence (SEQ ID NO. 5) consisting of an initiation codon, a His10 tag and a recognition site for HRV3C protease (Human rhinovirus C protease) was added at the N-terminus, so that the repeat region at positions 1 to 13 was increased by about 2-fold and translation was terminated at amino acid residue 1154. The amino acid sequence at the C-terminal end of the amino acid sequence shown in SEQ ID NO. 4 is identical to the amino acid sequence shown in SEQ ID NO. 3.

The modified fibroin of (1-i) can be composed of the amino acid sequence shown in SEQ ID NO. 4.

The 2 nd modified fibroin has an amino acid sequence whose domain sequence has a reduced content of glycine residues as compared to the naturally-derived fibroin. The modified fibroin of FIG. 2 can have an amino acid sequence comparable to that of a naturally-derived fibroin in which at least one or more glycine residues in REP have been replaced with other amino acid residues.

The modified fibroin of the 2 nd order may have an amino acid sequence corresponding to at least one or more of the motif sequences in which one glycine residue is replaced with another amino acid residue in at least one motif sequence selected from GGX and GPGXX (wherein G represents a glycine residue, P represents a proline residue, and X represents an amino acid residue other than glycine) in REP, as compared with a naturally-derived fibroin.

In the modified fibroin of item 2, the ratio of the motif sequence in which the glycine residue is replaced with another amino acid residue may be 10% or more with respect to the entire motif sequence.

The 2 nd modified fibroin comprises formula 1: the domain sequence shown in [ (A) _n motif-REP ] _m and may have the following amino acid sequence: when the total number of amino acid residues in the amino acid sequence consisting of XGX (wherein X represents an amino acid residue other than glycine) in the sequence obtained by removing the (a) _n motif located on the most C-terminal side from the domain sequence to the C-terminal side of the domain sequence is denoted as z, and the total number of amino acid residues in the sequence obtained by removing the (a) _n motif located on the most C-terminal side from the domain sequence to the C-terminal side is denoted as w, z/w is 30% or more, 40% or more, 50% or more, or 50.9% or more. (A) The number of alanine residues in the _n motif may be 83% or more, preferably 86% or more, more preferably 90% or more, still more preferably 95% or more, and still more preferably 100% (meaning that it is composed of only alanine residues) relative to the total number of amino acid residues.

The modified fibroin of item 2 preferably has an increased content of an amino acid sequence consisting of XGX by replacing one glycine residue in the GGX motif with another amino acid residue. In the modified fibroin of the 2 nd aspect, the content of the amino acid sequence composed of GGX in the domain sequence is preferably 30% or less, more preferably 20% or less, still more preferably 10% or less, still more preferably 6% or less, still more preferably 4% or less, and particularly preferably 2% or less. The content ratio of the amino acid sequence composed of GGX in the domain sequence can be calculated by the same method as the calculation method of the content ratio (z/w) of the amino acid sequence composed of XGX described below.

The method of calculating z/w is described in further detail. First, in the case where expression 1 is included: among the fibroin (modified fibroin or fibroin of natural origin) having the domain sequence shown in [ (A) _n motif-REP ] _m, the amino acid sequence consisting of XGX was extracted from the whole REP contained in the sequence obtained by removing the sequence from the most C-terminal side of (A) _n motif to the C-terminal side of the domain sequence. The total number of amino acid residues constituting XGX is z. For example, when 50 amino acid sequences consisting of XGX are extracted (no repetition), z is 50×3=150. In addition, for example, as in the case of an amino acid sequence consisting of XGXGX, when X (middle X) included in 2 pieces XGX is present, calculation is performed by subtracting a repetitive portion (5 amino acid residues in the case of XGXGX). w is the total number of amino acid residues contained in the sequence obtained by removing the sequence from the (A) _n motif located on the most C-terminal side to the C-terminal end of the domain sequence. For example, in the case of the domain sequence shown in fig. 1, w is 4+50+4+100+4+10+4+20+4+30=230 (except for the (a) _n motif located on the most C-terminal side). Then, by dividing z by w, z/w (%) can be calculated.

Here, the z/w in the fibroin of natural origin will be described. First, as described above, it was confirmed by a method of illustrating fibroin having amino acid sequence information registered in NCBI GenBank, and 663 kinds of fibroin (among them, 415 kinds of fibroin derived from spiders) were extracted. Among all the fibroin extracted, from the group comprising formula 1: the z/w was calculated according to the above calculation method in the amino acid sequence of a naturally-derived fibroin having a content of amino acid sequence composed of GGX in the fibroin of the domain sequence represented by [ (A) _n motif-REP ] _m ] of 6% or less. The results are shown in FIG. 2. The horizontal axis of fig. 2 represents z/w (%), and the vertical axis represents frequency. As can be seen from FIG. 2, the z/w in the naturally derived fibroin is less than 50.9% (up to 50.86%).

In the modified fibroin of the 2 nd, z/w is preferably 50.9% or more, more preferably 56.1% or more, further preferably 58.7% or more, further preferably 70% or more, further preferably 80% or more. The upper limit of z/w is not particularly limited, and may be, for example, 95% or less.

The 2 nd modified fibroin can be obtained, for example, by modifying as follows: at least a part of the nucleotide sequence encoding a glycine residue is substituted from the cloned gene sequence of a naturally-derived fibroin and other amino acid residues are encoded. In this case, as the modified glycine residue, one glycine residue in the GGX motif and the GPGXX motif may be selected, or the substitution may be performed so that the z/w ratio is 50.9% or more. Further, for example, it is also possible to design an amino acid sequence satisfying the above-described mode based on the amino acid sequence of a naturally-derived fibroin, and to obtain it by chemically synthesizing a nucleic acid encoding the designed amino acid sequence. In any case, in addition to the modifications corresponding to the substitution of glycine residues in REP with other amino acid residues in the amino acid sequence of the fibroin derived from natural sources, the modifications corresponding to the substitution, deletion, insertion and/or addition of one or more amino acid residues may be further performed.

The other amino acid residues are not particularly limited as long as they are amino acid residues other than glycine residues, but are preferably hydrophobic amino acid residues such as valine (V) residues, leucine (L) residues, isoleucine (I) residues, methionine (M) residues, proline (P) residues, phenylalanine (F) residues and tryptophan (W) residues, and hydrophilic amino acid residues such as glutamine (Q) residues, asparagine (N) residues, serine (S) residues, lysine (K) residues and glutamic acid (E) residues, more preferably valine (V) residues, leucine (L) residues, isoleucine (I) residues, phenylalanine (F) residues and glutamine (Q) residues, and further preferably glutamine (Q) residues.

More specific examples of the modified fibroin of the 2 nd item include modified fibroin having an amino acid sequence having 90% or more sequence identity with the amino acid sequence shown by (2-i) SEQ ID NO. 6 (Met-PRT 380), SEQ ID NO. 7 (Met-PRT 410), SEQ ID NO. 8 (Met-PRT 525) or SEQ ID NO. 9 (Met-PRT 799), or (2-ii) SEQ ID NO. 6, SEQ ID NO. 7, SEQ ID NO. 8 or SEQ ID NO. 9.

The modified fibroin of (2-i) is described. The amino acid sequence represented by SEQ ID NO. 6 is obtained by replacing all GGX in REP corresponding to the amino acid sequence represented by SEQ ID NO. 10 (Met-PRT 313) of a naturally occurring fibroin with GQX. The amino acid sequence shown in SEQ ID NO. 7 is obtained by deleting every 2 motifs of (A) _n from the N-terminal side to the C-terminal side of the amino acid sequence shown in SEQ ID NO. 6, and further inserting one [ (A) _n motif-REP ] just before the C-terminal sequence. The amino acid sequence shown in SEQ ID NO. 8 has 2 alanine residues inserted on the C-terminal side of each (A) _n motif of the amino acid sequence shown in SEQ ID NO. 7, and further has a serine (S) residue substituted for a part of the glutamine (Q) residue, and a part of the amino acid on the C-terminal side has been deleted so that it has almost the same molecular weight as SEQ ID NO. 7. The amino acid sequence shown in SEQ ID NO. 9 adds a predetermined hinge sequence and His tag sequence to the C-terminus of a sequence obtained by repeating a region of the 20 domain sequences present in the amino acid sequence shown in SEQ ID NO. 7 (in which several amino acid residues on the C-terminal side of the region are replaced) 4 times.

The value of z/w in the amino acid sequence shown in SEQ ID NO. 10 (corresponding to a naturally-derived fibroin) was 46.8%. The values of z/w in the amino acid sequence shown in SEQ ID NO.6, the amino acid sequence shown in SEQ ID NO. 7, the amino acid sequence shown in SEQ ID NO. 8 and the amino acid sequence shown in SEQ ID NO. 9 were 58.7%, 70.1%, 66.1% and 70.0%, respectively. The values of x/y of the amino acid sequences shown in SEQ ID Nos. 10, 6,7,8 and 9 (described below) were 15.0%, 93.4%, 92.7% and 89.8%, respectively, and the ratio of the saw teeth was 1:1.8 to 11.3.

The modified fibroin of (2-i) may be composed of the amino acid sequence shown in SEQ ID NO. 6, SEQ ID NO. 7, SEQ ID NO. 8 or SEQ ID NO. 9.

The modified fibroin of (2-ii) comprises an amino acid sequence having a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 6, SEQ ID NO. 7, SEQ ID NO. 8 or SEQ ID NO. 9. The engineered fibroin of (2-ii) is also of formula 1: protein of domain sequence shown in [ (A) _n motif-REP ] _m. The sequence identity is preferably 95% or more.

The modified fibroin of (2-ii) has a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 6, SEQ ID NO.7, SEQ ID NO. 8 or SEQ ID NO. 9, and when the total number of amino acid residues of an amino acid sequence consisting of XGX (wherein X represents an amino acid residue other than glycine) contained in REP is defined as z and the total number of amino acid residues of REP in the above-mentioned domain sequence is defined as w, z/w is preferably 50.9% or more.

The 2 nd engineered fibroin can comprise a tag sequence at either or both of the N-terminus and the C-terminus. Thus, the separation, fixation, detection, visualization and the like of the modified fibroin can be realized.

Examples of the tag sequence include affinity tags using specific affinity (binding property, affinity) with other molecules. Specific examples of the affinity tag include a histidine tag (His tag). His tags are short peptides comprising about 4 to 10 histidine residues, and have the property of specifically binding to metal ions such as nickel, and thus can be used for the isolation of modified fibroin by metal chelate chromatography (CHELATING METAL chromatography). Specific examples of the tag sequence include the amino acid sequence shown in SEQ ID NO. 11 (an amino acid sequence including a His tag sequence and a hinge sequence).

In addition, a tag sequence such as glutathione-S-transferase (GST) which specifically binds to glutathione or Maltose Binding Protein (MBP) which specifically binds to maltose can be used.

Furthermore, an "epitope tag" using an antigen-antibody reaction can be used. By adding a peptide (epitope) for exhibiting antigenicity as a tag sequence, an antibody against the epitope can be bound. Examples of the epitope tag include an HA (peptide sequence of influenza virus hemagglutinin) tag, myc tag, FLAG tag, and the like. By using epitope tags, the engineered fibroin can be easily purified with high specificity.

Further, a tag obtained by cleaving a tag sequence with a specific protease may be used. The modified fibroin after cleavage of the tag sequence can also be recovered by subjecting the protein adsorbed via the tag sequence to protease treatment.

More specific examples of modified fibroin containing a tag sequence include modified fibroin comprising an amino acid sequence having 90% or more sequence identity with an amino acid sequence represented by (2-iii) SEQ ID NO. 12 (PRT 380), SEQ ID NO. 13 (PRT 410), SEQ ID NO. 14 (PRT 525) or SEQ ID NO. 15 (PRT 799), or an amino acid sequence represented by (2-iv) SEQ ID NO. 12, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15.

The amino acid sequences shown in SEQ ID Nos. 16 (PRT 313), 12, 13, 14 and 15 are added with the amino acid sequence shown in SEQ ID No. 11 (including His tag sequence and hinge sequence) at the N-terminus of the amino acid sequences shown in SEQ ID Nos. 10, 6, 7, 8 and 9, respectively.

The modified fibroin of (2-iii) may be composed of the amino acid sequence shown in SEQ ID NO. 12, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15.

The modified fibroin of (2-iv) comprises an amino acid sequence having a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 12, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15. The engineered fibroin of (2-iv) is also comprised of formula 1: protein of domain sequence shown in [ (A) _n motif-REP ] _m. The sequence identity is preferably 95% or more.

The modified fibroin of (2-iv) has a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 12, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15, and when the total number of amino acid residues of an amino acid sequence consisting of XGX (wherein X represents an amino acid residue other than glycine) contained in REP is defined as z and the total number of amino acid residues of REP in the above-mentioned domain sequence is defined as w, z/w is preferably 50.9% or more.

The 2 nd modified fibroin can contain a secretion signal for releasing the protein produced by the recombinant protein production system to the outside of the host. The sequence of the secretion signal may be appropriately set according to the type of host.

The 3 rd engineered fibroin has an amino acid sequence with a reduced content of (a) _n motifs compared to the naturally-derived fibroin domain sequence. The domain sequence of the 3 rd engineered fibroin can have an amino acid sequence corresponding to a deletion of at least one or more of the (a) _n motifs as compared to the naturally-derived fibroin.

The 3 rd engineered fibroin can have an amino acid sequence corresponding to 10-40% of the (a) _n motif deleted from the naturally-derived fibroin.

The domain sequence of the 3 rd engineered fibroin may have an amino acid sequence corresponding to deletion of one (a) _n motif per 1 to 3 (a) _n motifs from at least the N-terminal side to the C-terminal side, compared with the naturally-derived fibroin.

The modified fibroin of the 3 rd domain may have an amino acid sequence corresponding to the sequential repetition of deletion of 2 consecutive (a) _n motifs and deletion of one (a) _n motif, at least from the N-terminal side to the C-terminal side, compared with the naturally-derived fibroin.

The domain sequence of the 3 rd engineered fibroin may have an amino acid sequence corresponding to the motif of (a) _n occurring once every 2 from the N-terminal side to the C-terminal side.

The 3 rd engineered fibroin comprises formula 1: the domain sequence shown in [ (A) _n motif-REP ] _m and may have the following amino acid sequence: comparing the number of amino acid residues of REP of adjacent 2 [ (A) _n motif-REP ] units sequentially from N-terminal side to C-terminal side, when the number of amino acid residues of REP with less number of amino acid residues is 1, and when the sum of the numbers of amino acid residues of adjacent 2 [ (A) _n motif-REP ] units with the ratio of the number of amino acid residues of the other REP being 1.8-11.3 is x, and when the total number of amino acid residues of the domain sequence is y, x/y is 20% or more, 30% or more, 40% or more, or 50% or more. (A) The number of alanine residues in the _n motif may be 83% or more, preferably 86% or more, more preferably 90% or more, still more preferably 95% or more, and still more preferably 100% (meaning that it is composed of only alanine residues) relative to the total number of amino acid residues.

The x/y calculation method is further described in detail with reference to FIG. 1. FIG. 1 shows the domain sequences after removal of the N-terminal and C-terminal sequences from the engineered fibroin. The domain sequence has, from the N-terminal side (left), the sequence of (A) _n th motif-1 REP (50 amino acid residues) - (A) _n th motif-2 REP (100 amino acid residues) - (A) _n th motif-3 REP (10 amino acid residues) - (A) _n th motif-4 REP (20 amino acid residues) - (A) _n th motif-5 REP (30 amino acid residues) - (A) _n motif.

The adjacent 2 [ (A) _n motif-REP ] units were selected in sequence from the N-terminal side to the C-terminal side in a non-repetitive manner. In this case, an unselected [ (A) _n motif-REP ] unit may be present. Fig. 1 shows pattern 1 (comparison of 1 st REP and 2 nd REP, and comparison of 3 rd REP and 4 th REP), pattern 2 (comparison of 1 st REP and 2 nd REP, and comparison of 4 th REP and 5 th REP), pattern 3 (comparison of 2 nd REP and 3 rd REP, and comparison of 4 th REP and 5 th REP), pattern 4 (comparison of 1 st REP and 2 nd REP). In addition, there are selection methods in addition.

Next, for each pattern, the amino acid residue numbers of each REP in the selected adjacent 2 [ (a) _n motif-REP ] units are compared. The comparison was performed by determining the ratio of the numbers of amino acid residues at the other side when the number of amino acid residues at the other side was 1. For example, in the case of comparing 1 st REP (50 amino acid residues) and 2 nd REP (100 amino acid residues), when 1 st REP having a relatively small number of amino acid residues is set to 1, the ratio of the number of amino acid residues of 2 nd REP is 100/50=2. Similarly, in the case of comparing 4 th REP (20 amino acid residues) and 5 th REP (30 amino acid residues), when 4 th REP having a relatively small number of amino acid residues is set to 1, the ratio of the number of amino acid residues of 5 th REP is 30/20=1.5.

In FIG. 1, the group of [ (A) _n motif-REP ] units whose ratio of the number of amino acid residues is 1.8 to 11.3 is shown by a solid line when one of the relatively smaller numbers of amino acid residues is 1. In this specification, this ratio is referred to as a saw tooth ratio. The group of [ (A) _n motif-REP ] units whose ratio of the number of amino acid residues is less than 1.8 or more than 11.3 when one of the relatively small numbers of amino acid residues is 1 is indicated by a dotted line.

In each mode, all amino acid residues of the adjacent 2 [ (a) _n motif-REP ] units shown in solid lines are added (not only REP, but also the amino acid residue of the (a) _n motif). The added total value is compared, and the total value of the mode in which the total value is largest (the maximum value of the total value) is set as x. In the example shown in fig. 1, the total value of pattern 1 is the largest.

Next, x/y (%) can be calculated by dividing x by the total number of amino acid residues y of the domain sequence.

In the modified fibroin of the 3 rd aspect, x/y is preferably 50% or more, more preferably 60% or more, further preferably 65% or more, still more preferably 70% or more, further preferably 75% or more, and particularly preferably 80% or more. The upper limit of x/y is not particularly limited, and may be, for example, 100% or less. When the serration ratio is 1:1.9 to 11.3, x/y is preferably 89.6% or more, when the serration ratio is 1:1.8 to 3.4, x/y is preferably 77.1% or more, when the serration ratio is 1:1.9 to 8.4, x/y is preferably 75.9% or more, and when the serration ratio is 1:1.9 to 4.1, x/y is preferably 64.2% or more.

When the 3 rd modified fibroin is a modified fibroin comprising at least 7 of the plurality of (a) _n motifs in the domain sequence and only alanine residues, x/y is preferably 46.4% or more, more preferably 50% or more, still more preferably 55% or more, still more preferably 60% or more, still more preferably 70% or more, and particularly preferably 80% or more. The upper limit of x/y is not particularly limited, and may be 100% or less.

Here, x/y in the fibroin of natural origin will be described. First, as described above, it was confirmed by a method of illustrating fibroin having amino acid sequence information registered in NCBI GenBank, and 663 kinds of fibroin (among them, 415 kinds of fibroin derived from spiders) were extracted. Of all the fibroin extracted, one from the group consisting of formula 1: in the amino acid sequence of the naturally-derived fibroin consisting of the domain sequence represented by [ (A) _n motif-REP ] _m ], x/y was calculated according to the above-described calculation method. FIG. 3 shows the results when the serration ratio is 1:1.9 to 4.1. The horizontal axis of fig. 3 represents x/y (%), and the vertical axis represents frequency. As can be seen from FIG. 3, the x/y in the fibroin of natural origin is less than 64.2% (maximum 64.14%).

For example, the 3 rd modified fibroin can be obtained by deleting one or more of the sequences encoding the motif of (a) _n from the cloned gene sequence of a naturally-derived fibroin so that x/y is 64.2% or more. Alternatively, it is also possible to design an amino acid sequence corresponding to a naturally-derived fibroin, to delete one or more (A) _n motifs so that x/y is 64.2% or more, and to chemically synthesize a nucleic acid encoding the designed amino acid sequence. In any case, in addition to the modifications corresponding to the deletion of the (a) _n motif in the amino acid sequence of the fibroin from natural sources, further modifications of the amino acid sequence corresponding to substitution, deletion, insertion and/or addition of one or more amino acid residues may be made.

More specific examples of the 3 rd modified fibroin include modified fibroin having an amino acid sequence having 90% or more sequence identity with the amino acid sequence shown by (3-i) SEQ ID NO. 17 (Met-PRT 399), SEQ ID NO.7 (Met-PRT 410), SEQ ID NO. 8 (Met-PRT 525) or SEQ ID NO. 9 (Met-PRT 799), or (3-ii) SEQ ID NO. 17, SEQ ID NO.7, SEQ ID NO. 8 or SEQ ID NO. 9.

The modified fibroin of (3-i) is described. The amino acid sequence shown in SEQ ID NO. 17 is obtained by deleting every 2 of the motif (A) _n from the N-terminal side to the C-terminal side in the amino acid sequence shown in SEQ ID NO. 10 (Met-PRT 313) corresponding to a naturally-derived fibroin, and further inserting one [ (A) _n motif-REP ] just before the C-terminal sequence. The amino acid sequence shown in SEQ ID NO. 7, SEQ ID NO. 8 or SEQ ID NO. 9 is as described in the modified fibroin of FIG. 2.

The serration ratio of the amino acid sequence shown in SEQ ID NO. 10 (corresponding to a naturally-derived fibroin) was 15.0% in the range of 1:1.8 to 11.3. The amino acid sequence shown in SEQ ID NO. 17 and the value of x/y in the amino acid sequence shown in SEQ ID NO. 7 were each 93.4%. The value of x/y in the amino acid sequence shown in SEQ ID NO. 8 was 92.7%. The value of x/y in the amino acid sequence shown in SEQ ID NO. 9 was 89.8%. The values of z/w in the amino acid sequences shown in SEQ ID Nos. 10, 17, 7, 8 and 9 were 46.8%, 56.2%, 70.1%, 66.1% and 70.0%, respectively.

The modified fibroin of (3-i) may be composed of the amino acid sequence shown in SEQ ID NO. 17, SEQ ID NO. 7, SEQ ID NO. 8 or SEQ ID NO. 9.

The modified fibroin of (3-ii) comprises an amino acid sequence having a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 17, SEQ ID NO. 7, SEQ ID NO. 8 or SEQ ID NO. 9. The engineered fibroin of (3-ii) is also of formula 1: protein of domain sequence shown in [ (A) _n motif-REP ] _m. The sequence identity is preferably 95% or more.

The modified fibroin of (3-ii) has a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 17, SEQ ID NO. 7, SEQ ID NO. 8 or SEQ ID NO. 9, and the maximum value of the total value obtained by adding the amino acid residues of adjacent 2 [ (A) _n motif-REP ] units, in which x/y is preferably 64.2% or more, is set to 1 when the number of amino acid residues of REP having a small number of amino acid residues is compared with the number of amino acid residues of REP of 2 [ (A) _n motif-REP ] units in order from the N-terminal side to the C-terminal side, and the ratio of the number of amino acid residues of another REP is 1.8 to 11.3 (the serration ratio is 1:1.8 to 11.3).

The 3 rd engineered fibroin can comprise the above-described tag sequences at either or both of the N-terminal and C-terminal ends.

More specific examples of modified fibroin containing a tag sequence include modified fibroin comprising an amino acid sequence having 90% or more sequence identity with an amino acid sequence represented by (3-iii) SEQ ID NO. 18 (PRT 399), SEQ ID NO. 13 (PRT 410), SEQ ID NO. 14 (PRT 525) or SEQ ID NO. 15 (PRT 799), or an amino acid sequence represented by (3-iv) SEQ ID NO. 18, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15.

The amino acid sequences shown in SEQ ID Nos. 18, 13, 14 and 15 are added with the amino acid sequence shown in SEQ ID No. 11 (including His tag sequence and hinge sequence) at the N-terminus of the amino acid sequences shown in SEQ ID Nos. 17, 7, 8 and 9, respectively.

The modified fibroin of (3-iii) may be composed of the amino acid sequence shown in SEQ ID NO. 18, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15.

The modified fibroin of (3-iv) comprises an amino acid sequence having a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 18, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15. The engineered fibroin of (3-iv) is also comprised of formula 1: protein of domain sequence shown in [ (A) _n motif-REP ] _m. The sequence identity is preferably 95% or more.

The modified fibroin of (3-iv) has a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 18, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15, and when the number of amino acid residues of REPs of adjacent 2 [ (A) _n motif-REP ] units is compared in order from the N-terminal side to the C-terminal side, the number of amino acid residues of REP having a small number of amino acid residues is 1, and the sum of the numbers of amino acid residues of adjacent 2 [ (A) _n motif-REP ] units having a ratio of the number of amino acid residues of the other REP of 1.8 to 11.3 is set to x, and when the total number of amino acid residues of the domain sequence is set to y, x/y is preferably 64.2% or more.

The 3 rd engineered fibroin can contain a secretion signal for releasing the protein produced by the recombinant protein production system to the outside of the host. The sequence of the secretion signal may be appropriately set according to the type of host.

The 4 th modified fibroin is a fibroin whose domain sequence has an amino acid sequence that reduces the content of glycine residues in addition to the content of (a) _n motif, as compared with a fibroin of natural origin. The domain sequence of the 4 th engineered fibroin can have an amino acid sequence corresponding to substitution of at least one or more glycine residues in REP with other amino acid residues in addition to deletion of at least one or more (a) _n motifs as compared to the naturally-derived fibroin. That is, the 4 th modified fibroin is a modified fibroin having the characteristics of both the 2 nd modified fibroin and the 3 rd modified fibroin described above. Specific embodiments and the like are as described in the 2 nd modified fibroin and the 3 rd modified fibroin.

More specific examples of the 4 th modified fibroin include modified fibroin having an amino acid sequence having a sequence identity of 90% or more with the amino acid sequence shown by (4-i) SEQ ID NO. 7 (Met-PRT 410), SEQ ID NO. 8 (Met-PRT 525), SEQ ID NO. 9 (Met-PRT 799), SEQ ID NO. 13 (PRT 410), SEQ ID NO. 14 (PRT 525) or SEQ ID NO. 15 (PRT 799), or (4-ii) SEQ ID NO. 7, SEQ ID NO. 8, SEQ ID NO. 9, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15. Specific embodiments of the modified fibroin comprising the amino acid sequences shown in SEQ ID NO. 7, SEQ ID NO. 8, SEQ ID NO. 9, SEQ ID NO. 13, SEQ ID NO. 14 or SEQ ID NO. 15 are as described above.

The modified fibroin of the 5th aspect may have an amino acid sequence which corresponds to a region in which one or more amino acid residues in REP have been replaced with an amino acid residue having a large hydrophobicity index and/or in which one or more amino acid residues having a large hydrophobicity index have been inserted into REP, as compared with a fibroin of natural origin.

The region with a large local hydrophobicity index is preferably composed of 2 to 4 consecutive amino acid residues.

The amino acid residue having a large hydrophobicity index is more preferably an amino acid residue selected from the group consisting of isoleucine (I), valine (V), leucine (L), phenylalanine (F), cysteine (C), methionine (M) and alanine (A).

The 5 th modified fibroin can be modified in comparison with a fibroin of natural origin by replacing one or more amino acid residues in REP with amino acid residues having a large hydrophobicity index and/or inserting one or more amino acid residues having a large hydrophobicity index into REP, and can be modified in comparison with a fibroin of natural origin by replacing, deleting, inserting and/or adding one or more amino acid residues.

For example, the 5 th modified fibroin can be obtained by replacing one or more hydrophilic amino acid residues (e.g., amino acid residues with negative hydrophobicity) in REP with hydrophobic amino acid residues (e.g., amino acid residues with positive hydrophobicity) and/or inserting one or more hydrophobic amino acid residues into REP from the cloned gene sequence of the naturally-derived fibroin. Alternatively, it is also possible to design an amino acid sequence corresponding to a naturally-derived fibroin, for example, by substituting one or more hydrophilic amino acid residues in REP with hydrophobic amino acid residues, and/or inserting one or more hydrophobic amino acid residues into REP, and chemically synthesizing a nucleic acid encoding the designed amino acid sequence. In any case, in addition to modifications corresponding to substitution of one or more hydrophilic amino acid residues in REP with hydrophobic amino acid residues in the amino acid sequence of a naturally derived fibroin and/or insertion of one or more hydrophobic amino acid residues in REP, modifications corresponding to substitution, deletion, insertion and/or addition of one or more amino acid residues may be further performed.

The 5 th modified fibroin comprises formula 1: the domain sequence shown in [ (A) _n motif-REP ] _m and may have the following amino acid sequence: in all REPs contained in the sequence obtained by removing the sequence from the C-terminal-most (A) _n motif from the domain sequence to the C-terminal end of the domain sequence, the total number of amino acid residues contained in the region where the average value of the hydrophobicity index of the continuous 4 amino acid residues is 2.6 or more is defined as p, and when the total number of amino acid residues contained in the sequence obtained by removing the sequence from the C-terminal-most (A) _n motif to the C-terminal end of the domain sequence is defined as q, p/q is 6.2% or more.

As the index of hydrophobicity of the amino acid residues, a known index (Hydropathy index:Kyte J,&Doolittle R(1982)"A simple method for displaying the hydropathic character of a protein",J.Mol.Biol.,157,pp.105-132)., specifically, a hydrophobicity index (hereinafter, also referred to as "HI") of each amino acid is used. ) As shown in table 1 below.

TABLE 1

Amino acids	HI	Amino acids	HI
				Isoleucine (Ile)	4.5	Tryptophan (Trp)	-0.9
Valine (Val)	4.2	Tyrosine (Tyr)	-1.3
				Leucine (Leu)	3.8	Proline (Pro)	-1.6
Phenylalanine (Phe)	2.8	Histidine (His)	-3.2
				Cysteine (Cys)	2.5	Asparagine (Asn)	-3.5
Methionine (Met)	1.9	Aspartic acid (Asp)	-3.5
				Alanine (Ala)	1.8	Glutamine (Gln)	-3.5
Glycine (Gly)	-0.4	Glutamic acid (Glu)	-3.5
				Threonine (Thr)	-0.7	Lysine (Lys)	-3.9
Serine (Ser)	-0.8	Arginine (Arg)	-4.5

The method of calculating p/q will be described in further detail. In the calculation, the following formula 1 is used: the sequence obtained by removing the sequence from the (A) _n motif located on the most C-terminal side to the C-terminal end of the domain sequence (hereinafter referred to as "sequence A") from the domain sequence represented by [ (A) _n motif-REP ] _m. First, the average value of the hydrophobicity index of consecutive 4 amino acid residues was calculated in all REPs included in the sequence a. The average value of the hydrophobicity index is obtained by dividing the sum of HI of each amino acid residue contained in the consecutive 4 amino acid residues by 4 (the number of amino acid residues). The average value of the hydrophobicity index was determined for all consecutive 4 amino acid residues (each amino acid residue was used for calculation of the average value 1 to 4 times). Next, a region in which the average value of the hydrophobicity index of the continuous 4 amino acid residues is 2.6 or more is determined. Even when a certain amino acid residue belongs to a plurality of "4 consecutive amino acid residues whose average value of hydrophobicity index is 2.6 or more", it is included as one amino acid residue in a region. Furthermore, the total number of amino acid residues contained in this region is p. In addition, the total number of amino acid residues contained in sequence a is q.

For example, when 20 "consecutive 4 amino acid residues whose average value of the hydrophobicity index is 2.6 or more" are extracted (no repetition), the region where the average value of the hydrophobicity index of the consecutive 4 amino acid residues is 2.6 or more contains 20 consecutive 4 amino acid residues (no repetition), and p is 20×4=80. For example, when "4 consecutive amino acid residues whose average value of the hydrophobicity index is 2.6 or more" at2 is repeated with only one amino acid residue, 7 amino acid residues are contained in a region whose average value of the hydrophobicity index of 4 consecutive amino acid residues is 2.6 or more (p=2×4-1=7. "-1" is the subtraction of the repeated portion). For example, in the case of the domain sequence shown in fig. 4, the "average value of hydrophobicity index is 2.6 or more, consecutive 4 amino acid residues" is not repeated and 7 are present, and thus p is 7×4=28. In addition, for example, in the case of the domain sequence shown in fig. 4, q is 4+50+4+40+4+10+4+20+4+30=170 (excluding the last (a) _n motif on the C-terminal side). Then, by dividing p by q, p/q (%) can be calculated. In the case of fig. 4, 28/170=16.47%.

In the modified fibroin of the 5 th aspect, p/q is preferably 6.2% or more, more preferably 7% or more, still more preferably 10% or more, still more preferably 20% or more, still more preferably 30% or more. The upper limit of p/q is not particularly limited, and may be 45% or less, for example.

For example, in the amino acid sequence of a cloned naturally-derived fibroin, the 5 th modified fibroin is obtained by substituting one or more hydrophilic amino acid residues (for example, amino acid residues with negative hydrophobicity) in REP with hydrophobic amino acid residues (for example, amino acid residues with positive hydrophobicity), and/or inserting one or more hydrophobic amino acid residues into REP so that the above-mentioned conditions of p/q are satisfied, and by modifying it to an amino acid sequence locally containing a region with a large hydrophobicity. Furthermore, for example, it is also possible to design an amino acid sequence satisfying the above conditions of p/q based on the amino acid sequence of a naturally-derived fibroin, and to obtain it by chemically synthesizing a nucleic acid encoding the designed amino acid sequence. In any case, in addition to the modification of replacing one or more amino acid residues in REP with amino acid residues having a large hydrophobicity index and/or inserting one or more amino acid residues in REP with a large hydrophobicity index, the modification of replacing, deleting, inserting and/or adding one or more amino acid residues may be further performed as compared with the naturally-derived fibroin.

The amino acid residues having a large hydrophobicity index are not particularly limited, but are preferably isoleucine (I), valine (V), leucine (L), phenylalanine (F), cysteine (C), methionine (M) and alanine (a), more preferably valine (V), leucine (L) and isoleucine (I).

More specific examples of the modified fibroin of item 5 include modified fibroin having an amino acid sequence having a sequence identity of 90% or more with the amino acid sequence represented by (5-i) SEQ ID NO. 19 (Met-PRT 720), SEQ ID NO. 20 (Met-PRT 665) or SEQ ID NO. 21 (Met-PRT 666), or (5-ii) SEQ ID NO. 19, SEQ ID NO. 20 or SEQ ID NO. 21.

The modified fibroin of (5-i) is described. The amino acid sequence shown in SEQ ID NO. 19 is an amino acid sequence (VLI) comprising 3 amino acid residues at every REP insertion 2, in addition to the domain sequence of the terminal on the C-terminal side, with respect to the amino acid sequence shown in SEQ ID NO. 7 (Met-PRT 410), and further comprises a part of glutamine (Q) residues replaced with serine (S) residues, and a part of the amino acid on the C-terminal side is deleted. The amino acid sequence shown in SEQ ID NO. 20 was inserted into an amino acid sequence (VLI) consisting of 3 amino acid residues at 1 every REP, relative to the amino acid sequence shown in SEQ ID NO. 8 (Met-PRT 525). The amino acid sequence shown in SEQ ID NO. 21 is inserted into an amino acid sequence (VLI) consisting of 3 amino acid residues at 2 positions every REP with respect to the amino acid sequence shown in SEQ ID NO. 8.

The modified fibroin of (5-i) may be composed of the amino acid sequence shown in SEQ ID NO. 19, SEQ ID NO. 20 or SEQ ID NO. 21.

The modified fibroin of (5-ii) comprises an amino acid sequence having a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 19, SEQ ID NO. 20 or SEQ ID NO. 21. The engineered fibroin of (5-ii) is also of formula 1: protein of domain sequence shown in [ (A) _n motif-REP ] _m. The sequence identity is preferably 95% or more.

The modified fibroin of (5-ii) has a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 19, SEQ ID NO. 20 or SEQ ID NO. 21, and preferably, when the total number of amino acid residues contained in a region where the average value of the hydrophobicity index of the consecutive 4 amino acid residues is 2.6 or more is p and the total number of amino acid residues contained in a sequence obtained by removing the sequence from the C-terminal side (A) _n motif to the C-terminal end of the domain sequence is q, p/q is 6.2% or more, from among REPs contained in the sequence obtained by removing the sequence from the C-terminal side (A) _n motif to the C-terminal end of the domain sequence from the domain sequence.

The 5 th engineered fibroin can comprise a tag sequence at either or both of the N-terminus and the C-terminus.

More specific examples of modified fibroin containing a tag sequence include modified fibroin having an amino acid sequence having 90% or more sequence identity with (5-iii) the amino acid sequence shown by SEQ ID No. 22 (PRT 720), SEQ ID No. 23 (PRT 665) or SEQ ID No. 24 (PRT 666), or (5-iv) the amino acid sequence shown by SEQ ID No. 22, SEQ ID No. 23 or SEQ ID No. 24.

The amino acid sequences shown in SEQ ID Nos. 22, 23 and 24 are obtained by adding the amino acid sequence shown in SEQ ID No. 11 (including His tag sequence and hinge sequence) to the N-terminal of the amino acid sequences shown in SEQ ID Nos. 19, 20 and 21, respectively.

The modified fibroin of (5-iii) may be composed of the amino acid sequence shown in SEQ ID No. 22, SEQ ID No. 23 or SEQ ID No. 24.

The modified fibroin of (5-iv) comprises an amino acid sequence having a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 22, SEQ ID NO. 23 or SEQ ID NO. 24. The engineered fibroin of (5-iv) is also comprised of formula 1: protein of domain sequence shown in [ (A) _n motif-REP ] _m. The sequence identity is preferably 95% or more.

The modified fibroin of (5-iv) has a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 22, SEQ ID NO. 23 or SEQ ID NO. 24, and preferably, when the total number of amino acid residues contained in a region where the average value of the hydrophobicity index of the consecutive 4 amino acid residues is 2.6 or more is p and the total number of amino acid residues contained in a sequence obtained by removing the sequence from the C-terminal side (A) _n motif to the C-terminal end of the domain sequence is q, p/q is 6.2% or more, from among REPs contained in the sequence obtained by removing the sequence from the C-terminal side (A) _n motif to the C-terminal end of the domain sequence from the domain sequence.

The 5 th modified fibroin can contain a secretion signal for releasing the protein produced by the recombinant protein production system to the outside of the host. The sequence of the secretion signal may be appropriately set according to the type of host.

The 6 th modified fibroin has an amino acid sequence with reduced glutamine residue content compared to a naturally derived fibroin.

The 6 th modified fibroin preferably contains at least 1 motif selected from the group consisting of GGX motif and GPGXX motif in the amino acid sequence of REP.

When the 6 th modified fibroin includes the GPGXX motif in REP, the GPGXX motif content is usually 1% or more, or may be 5% or more, and preferably 10% or more. The upper limit of the GPGXX motif content is not particularly limited, and may be 50% or less or 30% or less.

In the present specification, the "GPGXX motif content" is a value calculated by the following method.

In the case of the composition comprising formula 1: [ (a) _n motif-REP ] _m or formula 2: in the case of fibroin (modified fibroin or fibroin of natural origin) of the domain sequence shown in [ (A) _n motif-REP ] _m-(A)_n, in all REPs contained in the sequence obtained by removing the sequence from the domain sequence from the (A) _n motif located on the most C-terminal side to the C-terminal side of the domain sequence, when the total number of GPGXX motifs contained in the region multiplied by 3 (i.e., the total number of G and P in the GPGXX motif) is set as s, and the total number of amino acid residues of all REPs obtained by removing the sequence from the (A) _n motif located on the most C-terminal side to the C-terminal side of the domain sequence from the (A) _n motif is set as t, the GPGXX motif content is calculated as s/t.

In the calculation of the GPGXX motif content, "a sequence obtained by removing a sequence from the (a) _n motif located on the most C-terminal side to the C-terminal end of the domain sequence" was taken as a target in order to exclude the following effects: the "sequence from (A) _n motif located on the most C-terminal side to the C-terminal end of the domain sequence" (sequence corresponding to REP) may include a sequence having low correlation with a characteristic sequence of fibroin, and if m is small (that is, if the domain sequence is short), the calculation result of GPGXX motif content may be affected. In the case where the "GPGXX motif" is located at the C-terminus of REP, even when "XX" is "AA", for example, the "GPGXX motif" is treated as "GPGXX motif".

FIG. 5 is a schematic representation showing the domain sequence of an engineered fibroin. The method for calculating the GPGXX motif content will be specifically described with reference to fig. 5. First, in the domain sequence of the modified silk fibroin shown in fig. 5 ("[ (a) _n motif-REP ] _m-(A)_n motif" type), all REPs are contained in the "sequence obtained by removing the sequence from the domain sequence from the (a) _n motif located on the most C-terminal side to the C-terminal side of the domain sequence" (the sequence shown as "region a" in fig. 5), and thus the number of GPGXX motifs used for calculating s is 7,s to be 7x3=21. Similarly, since all REPs are contained in "the sequence obtained by removing the sequence from the (a) _n motif located on the most C-terminal side to the C-terminal side of the domain sequence" (the sequence shown as "region a" in fig. 5), the total number t of amino acid residues of all REPs obtained by further removing the (a) _n motif from the sequence is 50+40+10+20+30=150. Next, s/t (%) can be calculated by dividing s by t, which in the case of the engineered fibroin of fig. 5 is 21/150=14.0%.

In the modified fibroin of the 6 th aspect, the content of glutamine residue is preferably 9% or less, more preferably 7% or less, further preferably 4% or less, and particularly preferably 0%.

In the present specification, the "glutamine residue content" is a value calculated by the following method.

In the case of the composition comprising formula 1: [ (a) _n motif-REP ] _m or formula 2: in the case of fibroin (modified fibroin or fibroin of natural origin) having the domain sequence shown in the [ (a) _n motif-REP ] _m-(A)_n, the total number of glutamine residues contained in the region was u, and the total number of amino acid residues of all REPs obtained by removing the sequence from the (a) _n motif located on the most C-terminal side to the C-terminal side of the domain sequence and further removing the (a) _n motif was t, out of all REPs contained in the sequence obtained by removing the sequence from the domain sequence from the most C-terminal side to the C-terminal side of the domain sequence (corresponding to the sequence of "region a" in fig. 5), the glutamine residue content was calculated as u/t. The reason why the glutamine residue content is calculated is the same as that described above, with reference to "a sequence obtained by removing a sequence from the C-terminal most position of the (A) _n motif to the C-terminal of the domain sequence".

The domain sequence of the 6 th modified fibroin may have an amino acid sequence corresponding to deletion or substitution of one or more glutamine residues in REP to other amino acid residues, as compared with the naturally-derived fibroin.

The "other amino acid residue" may be any amino acid residue other than a glutamine residue, and is preferably an amino acid residue having a hydrophobicity index greater than that of a glutamine residue. The hydrophobicity index of the amino acid residues is shown in table 1.

As shown in table 1, as the amino acid residue having a hydrophobicity index larger than that of glutamine, there can be mentioned an amino acid residue selected from isoleucine (I), valine (V), leucine (L), phenylalanine (F), cysteine (C), methionine (M), alanine (a), glycine (G), threonine (T), serine (S), tryptophan (W), tyrosine (Y), proline (P) and histidine (H). Among these, amino acid residues selected from isoleucine (I), valine (V), leucine (L), phenylalanine (F), cysteine (C), methionine (M) and alanine (a) are more preferable, and amino acid residues selected from isoleucine (I), valine (V), leucine (L) and phenylalanine (F) are more preferable.

The degree of hydrophobicity of REP in the 6 th modified fibroin is preferably more than-0.8, more preferably-0.7 or more, still more preferably 0 or more, still more preferably 0.3 or more, and particularly preferably 0.4 or more. The upper limit of the degree of hydrophobicity of REP is not particularly limited, and may be 1.0 or less, or may be 0.7 or less.

In the present specification, the "degree of hydrophobicity of REP" is a value calculated by the following method.

In the case of the composition comprising formula 1: [ (a) _n motif-REP ] _m or formula 2: in the case of fibroin (modified fibroin or fibroin of natural origin) of the domain sequence shown in the [ (A) _n motif-REP ] _m-(A)_n motif, the total of all REPs contained in the sequence (corresponding to the sequence of "region A" in FIG. 5) obtained by removing the sequence from the C-terminal side of the (A) _n motif to the C-terminal side of the domain sequence from the domain sequence, was calculated as v/t when the total of the hydrophobicity index of each amino acid residue in the region was v, and the total of all REPs obtained by removing the sequence from the C-terminal side of the (A) _n motif to the C-terminal side of the domain sequence from the domain sequence and further removing the (A) _n motif was t. The reason why the "sequence obtained by removing the sequence from the C-terminal-most (A) _n motif to the C-terminal end of the domain sequence" is the same as the above reason in the calculation of the degree of hydrophobicity of REP.

The modified fibroin of the 6 th domain sequence may be modified to correspond to substitution, deletion, insertion and/or addition of one or more amino acid residues in REP, in addition to the modification corresponding to deletion of one or more glutamine residues in REP and/or substitution of one or more glutamine residues in REP with other amino acid residues, as compared with the naturally-derived fibroin.

For example, the 6 th modified fibroin can be obtained by deleting one or more glutamine residues in REP and/or replacing one or more glutamine residues in REP with other amino acid residues in the cloned gene sequence of the naturally-derived fibroin. Alternatively, it is also possible to design an amino acid sequence corresponding to a naturally-derived fibroin, to delete one or more glutamine residues in REP and/or to replace one or more glutamine residues in REP with other amino acid residues, and to chemically synthesize a nucleic acid encoding the designed amino acid sequence.

More specific examples of the 6 th modified fibroin include modified fibroin comprising an amino acid sequence represented by (6-i) SEQ ID NO. 25 (Met-PRT 888), SEQ ID NO. 26 (Met-PRT 965), SEQ ID NO. 27 (Met-PRT 889), SEQ ID NO. 28 (Met-PRT 916), SEQ ID NO. 29 (Met-PRT 918), SEQ ID NO. 30 (Met-PRT 699), SEQ ID NO. 31 (Met-PRT 698), SEQ ID NO. 32 (Met-PRT 966), SEQ ID NO. 41 (Met-PRT 917) or SEQ ID NO. 42 (Met-PRT 1028), and modified fibroin comprising an amino acid sequence having 90% or more sequence identity with the amino acid sequence represented by (6-ii) SEQ ID NO. 25, SEQ ID NO. 26, SEQ ID NO. 27, SEQ ID NO. 28, SEQ ID NO. 29, SEQ ID NO. 30, SEQ ID NO. 31, SEQ ID NO. 32, SEQ ID NO. 41 or SEQ ID NO. 42.

The modified fibroin of (6-i) is described. The amino acid sequence shown in SEQ ID NO. 25 is obtained by replacing all of QQQs in the amino acid sequence shown in SEQ ID NO. 7 (Met-PRT 410) with VL. The amino acid sequence represented by SEQ ID NO. 26 is obtained by replacing all of QQs in the amino acid sequence represented by SEQ ID NO. 7 with TS and replacing the remaining Qs with A. The amino acid sequence represented by SEQ ID NO. 27 is obtained by replacing all of QQs in the amino acid sequence represented by SEQ ID NO. 7 with VL and replacing the remaining Qs with I. The amino acid sequence represented by SEQ ID NO. 28 is obtained by replacing all of QQs in the amino acid sequence represented by SEQ ID NO. 7 with VI and replacing the remaining Qs with L. The amino acid sequence represented by SEQ ID NO. 29 is obtained by replacing all of QQs in the amino acid sequence represented by SEQ ID NO. 7 with VF and replacing the remaining Qs with I.

The amino acid sequence shown in SEQ ID NO. 30 is obtained by replacing all QQQs in the amino acid sequence shown in SEQ ID NO. 8 (Met-PRT 525) with VL. The amino acid sequence represented by SEQ ID NO. 31 is obtained by replacing all of QQs in the amino acid sequence represented by SEQ ID NO. 8 with VL and replacing the remaining Qs with I.

The amino acid sequence shown in SEQ ID NO. 32 is obtained by repeating the region of the 20 domain sequences present in the amino acid sequence shown in SEQ ID NO. 7 (Met-PRT 410) 2 times to obtain a sequence in which all QQs are replaced with VF and the remaining Qs are replaced with I.

The amino acid sequence represented by SEQ ID NO. 41 (Met-PRT 917) is an amino acid sequence obtained by replacing all of QQQs in the amino acid sequence represented by SEQ ID NO.7 with LI and replacing the remaining Qs with V. The amino acid sequence represented by SEQ ID NO. 42 (Met-PRT 1028) is an amino acid sequence obtained by replacing all of QQs in the amino acid sequence represented by SEQ ID NO.7 with IF and replacing the remaining Qs with T.

The amino acid sequences shown in SEQ ID Nos. 25, 26, 27, 28, 29, 30, 31, 32, 41 and 42 each have a glutamine residue content of 9% or less (Table 2).

TABLE 2

The modified fibroin of (6-i) may be composed of the amino acid sequences shown in SEQ ID NO.25, SEQ ID NO. 26, SEQ ID NO. 27, SEQ ID NO. 28, SEQ ID NO. 29, SEQ ID NO. 30, SEQ ID NO. 31, SEQ ID NO. 32, SEQ ID NO. 41 or SEQ ID NO. 42.

The modified fibroin of (6-ii) comprises an amino acid sequence having 90% or more sequence identity with the amino acid sequence represented by SEQ ID NO. 25, SEQ ID NO. 26, SEQ ID NO. 27, SEQ ID NO. 28, SEQ ID NO. 29, SEQ ID NO. 30, SEQ ID NO. 31, SEQ ID NO. 32, SEQ ID NO. 41 or SEQ ID NO. 42. The engineered fibroin of (6-ii) is also a modified fibroin comprising formula 1: [ (a) _n motif-REP ] _m, or formula 2: a protein having the domain sequence shown in the [ (A) _n motif REP ] _m-(A)_n motif. The sequence identity is preferably 95% or more.

The content of glutamine residue in the modified fibroin of (6-ii) is preferably 9% or less. The content of the GPGXX motif in the modified fibroin of (6-ii) is preferably 10% or more.

The 6 th engineered fibroin can comprise a tag sequence at either or both of the N-terminus and the C-terminus. Thus, the separation, fixation, detection, visualization and the like of the modified fibroin can be realized.

More specific examples of the 6 th modified fibroin having a tag sequence include modified fibroin comprising an amino acid sequence shown by (6-iii) SEQ ID NO. 33 (PRT 888), SEQ ID NO. 34 (PRT 965), SEQ ID NO. 35 (PRT 889), SEQ ID NO. 36 (PRT 916), SEQ ID NO. 37 (PRT 918), SEQ ID NO. 38 (PRT 699), SEQ ID NO. 39 (PRT 698), SEQ ID NO. 40 (PRT 966), SEQ ID NO. 43 (PRT 917) or SEQ ID NO. 44 (PRT 1028), and modified fibroin comprising an amino acid sequence having 90% or more sequence identity with the amino acid sequence shown by (6-iv) SEQ ID NO. 33, SEQ ID NO. 34, SEQ ID NO. 35, SEQ ID NO. 36, SEQ ID NO. 37, SEQ ID NO. 38, SEQ ID NO. 39, SEQ ID NO. 40, SEQ ID NO. 43 or SEQ ID NO. 44.

The amino acid sequences shown in SEQ ID Nos. 33, 34, 35, 36, 37, 38, 39, 40, 43 and 44 are added with the amino acid sequence shown in SEQ ID No. 11 (including His tag sequence and hinge sequence) at the N-terminus of the amino acid sequences shown in SEQ ID Nos. 25, 26, 27, 28, 29, 30, 31, 32, 41 and 42, respectively. Since the tag sequence was added only to the N-terminus, the glutamine residue content was not changed, and the glutamine residue content of the amino acid sequences shown in SEQ ID Nos. 33, 34, 35, 36, 37, 38, 39, 40, 43 and 44 were all 9% or less (Table 3).

TABLE 3

The modified fibroin of (6-iii) may be composed of the amino acid sequence shown as SEQ ID NO. 33, SEQ ID NO. 34, SEQ ID NO. 35, SEQ ID NO. 36, SEQ ID NO. 37, SEQ ID NO. 38, SEQ ID NO. 39, SEQ ID NO. 40, SEQ ID NO. 43 or SEQ ID NO. 44.

The modified fibroin of (6-iv) comprises an amino acid sequence having a sequence identity of 90% or more with the amino acid sequence represented by SEQ ID NO. 33, SEQ ID NO. 34, SEQ ID NO. 35, SEQ ID NO. 36, SEQ ID NO. 37, SEQ ID NO. 38, SEQ ID NO. 39, SEQ ID NO. 40, SEQ ID NO. 43 or SEQ ID NO. 44. The modified fibroin of (6-iv) is also a modified fibroin comprising formula 1: [ (a) _n motif-REP ] _m, or formula 2: a protein having the domain sequence shown in the [ (A) _n motif REP ] _m-(A)_n motif. The sequence identity is preferably 95% or more.

The content of glutamine residue in the modified fibroin of (6-iv) is preferably 9% or less. The content of the GPGXX motif in the modified fibroin of (6-iv) is preferably 10% or more.

The 6 th modified fibroin can contain a secretion signal for releasing the protein produced by the recombinant protein production system to the outside of the host. The sequence of the secretion signal may be appropriately set according to the type of host.

The modified fibroin may be modified fibroin having at least 2 or more of the features of modified fibroin 1, modified fibroin 2, modified fibroin 3, modified fibroin 4, modified fibroin 5 and modified fibroin 6.

The modified fibroin can be hydrophilic modified fibroin or hydrophobic modified fibroin. The hydrophobic modified fibroin means a modified fibroin in which the sum of the index of Hydrophobicity (HI) of all amino acid residues constituting the modified fibroin is calculated, and then the sum is divided by the total number of amino acid residues to obtain a value (average HI) of more than-0.8. More preferably, the modified fibroin has an average HI of-0.6 or more, still more preferably an average HI of-0.4 or more, still more preferably an average HI of-0.2 or more, and particularly preferably an average HI of 0 or more. The hydrophobicity index is shown in table 1. The hydrophilic spider silk protein is modified silk fibroin having an average HI of-0.8 or less. The modified fibroin of the present embodiment preferably has an average hydrophobicity index of greater than-0.8, or preferably-0.7 or more, or preferably-0.6 or more, more preferably-0.5 or more, or preferably-0.4 or more, or preferably-0.3 or more, or preferably-0.2 or more, or preferably-0.1 or more, more preferably 0 or more, or more preferably 0.1 or more, or more preferably 0.2 or more, still more preferably 0.3 or more, and particularly preferably 0.4 or more.

Examples of the hydrophobic modified fibroin include the 6 th modified fibroin described above. More specific examples of the hydrophobically modified fibroin include modified fibroin comprising the amino acid sequence shown by SEQ ID NO. 27, SEQ ID NO. 28, SEQ ID NO. 29, SEQ ID NO. 30, SEQ ID NO. 31, SEQ ID NO. 32, SEQ ID NO. 33 or SEQ ID NO. 43, and the amino acid sequence shown by SEQ ID NO. 35, SEQ ID NO. 37, SEQ ID NO. 38, SEQ ID NO. 39, SEQ ID NO. 40, SEQ ID NO. 41 or SEQ ID NO. 44.

Examples of the modified hydrophilic fibroin include the 1 st modified fibroin, the 2 nd modified fibroin, the 3 rd modified fibroin, the 4 th modified fibroin and the 5 th modified fibroin. More specific examples of the modified hydrophilic fibroin include an amino acid sequence represented by SEQ ID NO. 4, an amino acid sequence represented by SEQ ID NO. 6, SEQ ID NO. 7, SEQ ID NO. 8 or SEQ ID NO. 9, an amino acid sequence represented by SEQ ID NO. 13, SEQ ID NO. 12, SEQ ID NO. 14 or SEQ ID NO. 15, an amino acid sequence represented by SEQ ID NO. 18, SEQ ID NO. 7, SEQ ID NO. 8 or SEQ ID NO. 9, an amino acid sequence represented by SEQ ID NO. 17, SEQ ID NO. 12, SEQ ID NO. 14 or SEQ ID NO. 15, and an amino acid sequence represented by SEQ ID NO. 19, SEQ ID NO. 20 or SEQ ID NO. 21.

The modified fibroin according to the present embodiment may be a fibroin containing only 1 modified fibroin, or a fibroin containing 2 or more modified fibroin in combination. Further, it may be a fibroin that is combined and contains an altered fibroin and a structural protein other than the altered fibroin.

The structural protein may be a polypeptide derived from the above-mentioned natural structural protein, i.e., a recombinant polypeptide.

As the structural protein derived from collagen, for example, a protein comprising formula 3: the protein having the domain sequence represented by [ REP2] _p (wherein, in formula 3, p represents an integer of 5 to 300. REP2 represents an amino acid sequence composed of Gly-X-Y, X and Y represent arbitrary amino acid residues other than Gly. The plurality of REP 2's may be identical to each other or different from each other. Specifically, a protein comprising the amino acid sequence represented by SEQ ID NO. 45. Specifically, the amino acid sequence represented by SEQ ID NO. 45 is obtained by adding the amino acid sequence represented by SEQ ID NO. 11 (tag sequence and hinge sequence) to the N-terminus of the amino acid sequence from the 301 th residue to the 540 th residue corresponding to the repetitive part and motif of human type IV collagen obtained from NCBI database (GenBank accession numbers: CAA56335.1, GI: 3702452).

As the structural protein derived from the leg elastic protein, for example, a protein comprising formula 4: in particular, the amino acid sequence represented by SEQ ID NO. 46 is a protein obtained by adding an amino acid sequence represented by SEQ ID NO. 49 (tag sequence and hinge sequence) to the amino acid sequence represented by SEQ ID NO. 19 in the amino acid sequence represented by SEQ ID NO. 46 (GenBank accession numbers NP 611157, gl:24654243 of NCBI) such that the Thr at the 87 th residue is replaced with Ser, and the amino acid sequence at the 95 th residue is replaced with the N-terminal residue at the end of the amino acid sequence represented by SEQ ID NO. 19.

Examples of the structural protein derived from elastin include proteins having an amino acid sequence such as NCBI GenBank accession No. AAC98395 (human), I47076 (sheep), NP786966 (bovine). Specifically, a protein comprising the amino acid sequence shown in SEQ ID NO. 47 is exemplified. The amino acid sequence shown in SEQ ID NO. 47 adds the amino acid sequence shown in SEQ ID NO. 11 (tag sequence and hinge sequence) to the N-terminus of the amino acid sequence from 121 th residue to 390 th residue of the amino acid sequence of GenBank accession No. AAC98395 of NCBI.

Examples of the structural protein derived from keratin include goat (Capra hircus) type I keratin. Specifically, a protein including an amino acid sequence shown in SEQ ID NO. 48 (an amino acid sequence of GenBank accession No. ACY30466 of NCBI) is exemplified.

(Method for producing recombinant structural protein)

For example, the recombinant structural protein according to any of the above embodiments may also be produced by expressing a nucleic acid sequence encoding the recombinant structural protein by transforming a host with an expression vector having one or more regulatory sequences operably linked to the nucleic acid sequence. In the following, modified fibroin will be described as an example.

The method for producing the nucleic acid encoding the modified fibroin is not particularly limited. For example, the nucleic acid can be produced by using a gene encoding a natural fibroin, amplifying and cloning the gene by Polymerase Chain Reaction (PCR) or the like, and then modifying the gene by genetic engineering or chemical synthesis. The method of chemically synthesizing nucleic acid is not particularly limited, and for example, a gene can be chemically synthesized by ligating oligonucleotides which are automatically synthesized by AKTA oligopilot plus/100 (GE HEALTHCARE. Japan Co., ltd.) based on amino acid sequence information of fibroin obtained from NCBI network database or the like, using PCR or the like. In this case, in order to facilitate purification and/or confirmation of the modified fibroin, a nucleic acid encoding the modified fibroin composed of an amino acid sequence in which an amino acid sequence composed of an initiation codon and a His10 tag is added to the N-terminus of the amino acid sequence may be synthesized.

Regulatory sequences are sequences (e.g., promoters, enhancers, ribosome binding sequences, transcription termination sequences, etc.) that control the expression of an engineered fibroin in a host, and can be appropriately selected depending on the type of host. As the promoter, an inducible promoter that functions in a host cell and can express and induce the modified fibroin can be used. An inducible promoter is a promoter that can control transcription by the presence of an inducing substance (expression inducer), the absence of a repressor, or a physical factor such as an increase or decrease in temperature, osmotic pressure, or pH.

The type of the expression vector may be appropriately selected from a plasmid vector, a viral vector, a cosmid vector, a fossild vector, an artificial chromosome vector, and the like, depending on the type of the host. As the expression vector, a vector which is capable of autonomous replication in a host cell or is capable of integrating into a chromosome of a host and which contains a promoter at a position where a nucleic acid encoding an altered fibroin can be transcribed is preferably used.

As the host, any of prokaryotes, and eukaryotes such as yeast, filamentous fungi, insect cells, animal cells, and plant cells can be suitably used.

Preferred examples of the prokaryotic host include bacteria belonging to the genus Escherichia, brevibacterium, serratia, bacillus, microbacterium, brevibacterium, corynebacterium, pseudomonas, and the like. Examples of the microorganism belonging to the genus Escherichia include Escherichia coli. Examples of the microorganism belonging to the genus Brevibacillus include Brevibacillus georginata. Examples of microorganisms belonging to the genus Serratia include Serratia liquefaciens. Examples of the microorganism belonging to the genus Bacillus include Bacillus subtilis. Examples of the microorganism belonging to the genus Microbacterium include Microbacterium ammoniagenes. Examples of the microorganism belonging to the genus Brevibacterium include Brevibacterium furcate. Examples of the microorganism belonging to the genus Corynebacterium include Corynebacterium ammoniagenes. Examples of the microorganism belonging to the genus Pseudomonas include Pseudomonas putida.

In the case of using a prokaryote as a host, examples of vectors into which a nucleic acid encoding an altered fibroin is introduced include pBTrp (manufactured by Boringer John), pGEX (manufactured by Pharmacia), pUC18, pBluescriptII, pSupex, pET22b, pCold, pUB110, pNCO2 (Japanese patent application laid-open No. 2002-238569), and the like.

Examples of eukaryotic hosts include yeasts and filamentous fungi (such as molds). Examples of yeasts include yeasts belonging to the genus Saccharomyces, pichia, schizosaccharomyces, and the like. Examples of the filamentous fungi include filamentous fungi belonging to the genus Aspergillus, penicillium, trichoderma (Trichoderma), and the like.

In the case of eukaryotic organisms as hosts, examples of vectors into which nucleic acids encoding modified fibroin are introduced include YEP13 (ATCC 37115) and YEP24 (ATCC 37051). As a method for introducing the expression vector into the host cell, any method may be used as long as DNA is introduced into the host cell. Examples of the method using calcium ions include Proc. Natl. Acad. Sci. USA,69,2110 (1972), electroporation, spheroplast, protoplast, lithium acetate, and competent methods.

As a method of expressing nucleic acids based on a host transformed with an expression vector, secretory production, fusion protein expression, and the like can be performed according to the method described in molecular cloning, 2 nd edition, in addition to direct expression.

The modified fibroin can be produced, for example, by culturing a host transformed with an expression vector in a culture medium, producing and accumulating the modified fibroin in the culture medium, and collecting the modified fibroin from the culture medium. The method of culturing the host in the culture medium may be carried out according to a method generally used for culturing the host.

When the host is a prokaryote such as Escherichia coli or a eukaryote such as yeast, any of natural medium and synthetic medium may be used as the medium for culture as long as the medium contains a carbon source, a nitrogen source, an inorganic salt, and the like which can be assimilated by the host and can efficiently culture the host.

As the carbon source, any substance which can be assimilated by the transformed microorganism may be used, and for example, glucose, fructose, sucrose, and molasses containing these sugars, carbohydrates such as starch and starch hydrolysates, organic acids such as acetic acid and propionic acid, and alcohols such as ethanol and propanol may be used. Examples of the nitrogen source include ammonium salts of inorganic or organic acids such as ammonia, ammonium chloride, ammonium sulfate, ammonium acetate and ammonium phosphate, other nitrogen-containing compounds, peptone, meat extract, yeast extract, corn steep liquor, casein hydrolysate, soybean meal and soybean meal hydrolysate, and various fermentation tubes and their digested products. Examples of the inorganic salts include potassium hydrogen phosphate, dipotassium hydrogen phosphate, magnesium sulfate, sodium chloride, ferrous sulfate, manganese sulfate, copper sulfate, and calcium carbonate.

The culture of a prokaryote such as E.coli or a eukaryote such as yeast can be carried out under aerobic conditions such as shaking culture or deep aeration agitation culture. The culture temperature is, for example, 15 to 40 ℃. The incubation time is usually 16 hours to 7 days. The pH of the culture medium during the cultivation is preferably maintained between 3.0 and 9.0. The pH of the culture medium can be adjusted using an inorganic acid, an organic acid, an alkali solution, urea, calcium carbonate, ammonia, or the like.

In addition, antibiotics such as ampicillin and tetracycline may be added to the medium for culture as needed. When culturing a microorganism transformed by using an inducible promoter as an expression vector of the promoter, an inducer may be added to the medium as required. For example, when culturing a microorganism transformed with an expression vector using the lac promoter, isopropyl-. Beta. -D-thiogalactopyranoside and the like may be added to the medium; in the case of culturing a microorganism transformed with an expression vector using trp promoter, indoleacrylic acid or the like may be added to the medium.

Isolation and purification of the expressed engineered fibroin can be performed by a usual method. For example, in the case where the modified fibroin is expressed in cells in a dissolved state, after completion of the culture, the host cells are recovered by centrifugation, suspended in an aqueous buffer, and then the host cells are crushed by an ultrasonic crusher, a French press, a Manton-Gaulin homogenizer, a Dyno-MILL (DYNO-MILL), or the like to obtain a cell-free extract. From the supernatant obtained by subjecting the cell-free extract to centrifugal separation, purified preparations can be obtained by a method commonly used for separation and purification of proteins, namely, a solvent extraction method, a salting-out method using ammonium sulfate or the like, a desalting method, a precipitation method using an organic solvent, an anion exchange chromatography method using a resin such as Diethylaminoethyl (DEAE) -agarose, DIAION HPA-75 (manufactured by mitsubishi chemical company), a cation exchange chromatography method using a resin such as S-Sepharose FF (manufactured by Pharmacia), a hydrophobic chromatography method using a resin such as butyl agarose or phenyl agarose, a gel filtration method using a molecular sieve, an affinity chromatography, a chromatofocusing method, an isoelectric point electrophoresis method, or the like, alone or in combination.

In the case where the modified fibroin is expressed as an insoluble substance in cells, the insoluble substance of the modified fibroin is recovered as a precipitated fraction by disrupting and centrifuging the host cells after they are recovered. The insoluble body of recovered modified fibroin can be solubilized using a protein denaturing agent. After this procedure, a purified preparation of modified fibroin was obtained by the same separation and purification method as described above. In the case where the modified fibroin is secreted outside the cells, the modified fibroin can be recovered from the culture supernatant. That is, a culture supernatant is obtained by treating a culture by a method such as centrifugation, and a purified preparation can be obtained from the culture supernatant by using the same separation and purification method as described above.

(Spinning dope)

The spinning dope (dope) according to the present embodiment includes a recombinant structural protein (for example, modified fibroin) and a solvent. Next, a spinning dope containing modified fibroin will be described as an example of a recombinant structural protein.

The solvent of the spinning dope according to the present embodiment may be any solvent that can dissolve the modified fibroin, and examples thereof include organic solvents. Examples of the organic solvent include Hexafluoroisopropanol (HFIP), hexafluoroacetone (HFA), dimethyl sulfoxide (DMSO), N-Dimethylformamide (DMF), N-Dimethylacetamide (DMA), 1, 3-dimethyl-2-imidazolidinone (DMI), N-methyl-2-pyrrolidone (NMP), acetonitrile, N-methylmorpholine N-oxide (NMO), and formic acid. From the viewpoint of better solubility of the modified fibroin, HFIP, DMSO and formic acid are more preferable, and DMSO and formic acid are further preferable. These organic solvents may also contain water. These solvents may be used singly or in combination of 1 or more than 2.

The concentration of modified fibroin in the dope according to the present embodiment is preferably 10 to 50 wt%, more preferably 10 to 40 wt%, more preferably 15 to 35 wt%, still more preferably 20 to 35 wt%, more preferably 25 to 35 wt%, still more preferably 27 to 33 wt%, and particularly preferably 28 to 32 wt%, when the total amount of the dope is 100 wt%. When the concentration of the modified fibroin is 10 wt% or more, the productivity is further improved. When the concentration of the modified fibroin is 50 wt% or less, the spinning dope can be ejected from the spinning nozzle more stably, and productivity can be improved.

To the dope according to the present embodiment, an inorganic salt may be added as necessary. The inorganic salt may function as a dissolution promoter for the modified fibroin. Examples of the inorganic salt include alkali metal halides, alkaline earth metal halides, and alkaline earth metal nitrates. Specific examples of the inorganic salt include lithium carbonate, lithium chloride, calcium nitrate, lithium bromide, barium bromide, calcium bromide, barium chlorate, sodium perchlorate, lithium perchlorate, barium perchlorate, calcium perchlorate, and magnesium perchlorate. At least 1 inorganic salt thereof may also be added to the solvent.

The method for producing the spinning dope according to the present embodiment is not particularly limited, and the modified fibroin and the solvent may be mixed in any order. To facilitate dissolution, the dope may be stirred or shaken for a period of time. In this case, the spinning dope may be heated to a temperature at which it is soluble, depending on the modified fibroin and solvent used, if necessary. The spinning dope may be heated to 30℃or higher, 40℃or higher, 50℃or higher, 60℃or higher, 70℃or higher, 80℃or higher, or 90℃or higher, for example. From the viewpoint of further preventing the decomposition of the modified fibroin, it is preferably 40 ℃. The upper limit of the heating temperature is, for example, not more than the boiling point of the solvent.

The viscosity of the spinning dope according to the present embodiment can be appropriately set according to the use of the fiber, the spinning method, and the like. For example, the temperature may be 60,000 to 130,000 mPas sec or 65,000 to 125,000 mPas sec at 20 ℃. For example, 500 to 35,000 mPas, 1,000 to 35,000 mPas, 3,000 to 30,000 mPas, 500 to 20,000 mPas, 500 to 15,000 mPas, 1,000 to 12,000 mPas, 1,500 to 10,000 mPas, 1,500 to 8,000 mPas, and the like can be used at 35 ℃. Further, for example, 500 to 35,000 mPas, 1,000 to 35,000 mPas, 5,000 to 35,000 mPas, 10,000 to 30,000 mPas, 5,000 to 20,000 mPas, 8,000 to 20,000 mPas, 9,000 to 18,000 mPas, 9,000 to 16,000 mPas, 10,000 to 15,000 mPas, 12,000 to 30,000 mPas, 12,000 to 28,000 mPas, 12,000 to 18,000 mPas, 12,000 to 16,000 mPas, or the like at 40 ℃. The viscosity of the dope may be measured using, for example, an "EMS viscometer" trade name manufactured by Kyoto electronic industries, inc.

[ Coagulating liquid ]

The coagulation liquid according to the present embodiment may be any solvent that can be desolvated, and examples thereof include lower alcohols having 1 to 5 carbon atoms such as methanol, ethanol, and 2-propanol, ketones such as acetone, water, and aqueous solutions having a PH of 0.25 or more and a PH of 10.00 or less. The above solvents may be appropriately combined and used as a mixed solvent.

The coagulation liquid according to one embodiment mainly contains water or an aqueous solution having a pH of 0.25 or more and a pH of 10.00 or less. Thus, a method for producing a protein fiber can be provided, which reduces production costs and environmental load. The aqueous solution may be a saline solution, an aqueous acid solution, or a mixed solution of a saline solution and an aqueous acid solution, or a saline solution. The mixed solution of the aqueous salt solution and the aqueous acid solution is not limited to the solution in which the aqueous salt solution and the aqueous acid solution are mixed, but includes a solution in which the aqueous salt solution is mixed with the aqueous acid solution, and a solution in which the aqueous salt solution and the aqueous acid solution are dissolved.

(Acid aqueous solution)

The aqueous acid solution may be an aqueous solution of carboxylic acid or the like, and specific examples of carboxylic acid include formic acid, acetic acid, propionic acid, citric acid, oxalic acid, and the like. These solvents may be used alone or in combination of 2 or more. For example, the aqueous acid solution may be an aqueous citric acid solution or an aqueous formic acid solution.

(Saline solution)

The aqueous salt solution includes an aqueous salt solution of an organic salt or an inorganic salt, and a mixed aqueous solution of an organic salt and an inorganic salt.

Examples of the organic salt include carboxylate salts, and specific examples of the carboxylate salts include formate salts, acetate salts, propionate salts, citrate salts, and oxalate salts. For example, the organic salts may also be formate, acetate and citrate.

Specific examples of formate include ammonium formate, potassium formate, sodium formate, lithium formate, magnesium formate, and calcium formate.

Specific examples of the acetate salt include ammonium acetate, potassium acetate, sodium acetate, lithium acetate, magnesium acetate, and calcium acetate.

Specific examples of the propionate include ammonium propionate, potassium propionate, sodium propionate, lithium propionate, magnesium propionate, and calcium propionate.

Specific examples of the citrate include ammonium citrate, potassium citrate, sodium citrate, lithium citrate, magnesium citrate, and calcium citrate. For example, the citrate may include at least 1 selected from the group consisting of ammonium citrate, potassium citrate, sodium citrate, magnesium citrate, and calcium citrate, may further include at least 1 selected from the group consisting of ammonium citrate, potassium citrate, and sodium citrate, may further include at least 1 selected from the group consisting of potassium citrate and sodium citrate, and may further be sodium citrate.

Specific examples of the oxalate include ammonium oxalate, potassium oxalate, sodium oxalate, lithium oxalate, magnesium oxalate, calcium oxalate, and the like. The carboxylate is more preferably sodium carboxylate, and specific examples of sodium carboxylate include sodium formate, sodium acetate, sodium propionate, sodium oxalate, and the like.

Specific examples of the inorganic salt include normal salts, acidic salts, and basic salts.

Specific examples of the normal salt include sulfate, chloride, nitrate, iodide salt, thiocyanate, and carbonate.

Specific examples of the sulfate include ammonium sulfate, potassium sulfate, sodium sulfate, lithium sulfate, magnesium sulfate, and calcium sulfate. For example, the sulfate may include at least 1 selected from the group consisting of ammonium sulfate, sodium sulfate, magnesium sulfate, and calcium sulfate, may include at least 1 selected from the group consisting of ammonium sulfate and sodium sulfate, and may be sodium sulfate.

Specific examples of the chloride include ammonium chloride, potassium chloride, sodium chloride, lithium chloride, magnesium chloride, and calcium chloride. For example, the chloride may include at least 1 selected from the group consisting of ammonium chloride, potassium chloride, sodium chloride, lithium chloride, calcium chloride, and magnesium chloride, and the chloride may further include at least 1 selected from the group consisting of potassium chloride, sodium chloride, and calcium chloride, and may further include at least 1 selected from the group consisting of sodium chloride and calcium chloride, and may also be sodium chloride.

Specific examples of the nitrate include ammonium nitrate, potassium nitrate, sodium nitrate, lithium nitrate, magnesium nitrate, and calcium nitrate.

Specific examples of the iodide salt include ammonium iodide, potassium iodide, sodium iodide, lithium iodide, magnesium iodide, and calcium iodide.

Specific examples of thiocyanate include ammonium thiocyanate, potassium thiocyanate, sodium thiocyanate, lithium thiocyanate, magnesium thiocyanate, calcium thiocyanate, and guanidine thiocyanate.

Specific examples of the carbonate include ammonium carbonate, potassium carbonate, sodium carbonate, lithium carbonate, magnesium carbonate, and calcium carbonate.

Specific examples of the acidic salt include hydrogen sulfate, hydrogen phosphate, and hydrogen carbonate.

Specific examples of the bisulfate include ammonium bisulfate, potassium bisulfate, sodium bisulfate, lithium bisulfate, magnesium bisulfate, calcium bisulfate and the like.

Specific examples of the hydrogen phosphate include sodium dihydrogen phosphate, disodium hydrogen phosphate, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, ammonium dihydrogen phosphate, diammonium hydrogen phosphate, magnesium dihydrogen phosphate, calcium dihydrogen phosphate, and dicalcium hydrogen phosphate.

Specific examples of the bicarbonate include ammonium bicarbonate, potassium bicarbonate, sodium bicarbonate, lithium bicarbonate, magnesium bicarbonate, and calcium bicarbonate.

Specific examples of the alkali salt include calcium chloride and magnesium chloride.

The acid, acid aqueous solution, salt and brine solution may be used alone or in combination of 1 or more than 2.

Examples of the salt mixed aqueous solution in which 2 or more salts or brine solutions are mixed include a mixed aqueous solution of the organic salt, a mixed aqueous solution of the inorganic salt, a mixed aqueous solution of the organic salt and the inorganic salt, and the like, and brackish water and sea water are particularly preferable from the viewpoint of reducing the production cost. Brackish water and seawater are known to include mainly potassium chloride, sodium chloride, magnesium sulfate and calcium sulfate.

The coagulating liquid preferably contains a saline solution, more preferably a saline solution. By containing a salt, the desolvation rate can be further improved. The salt more preferably includes at least 1 of the group consisting of carboxylate, sulfate, chloride, hydrogen phosphate and bicarbonate, still more preferably includes at least 1 of the group consisting of carboxylate, sulfate and chloride, still more preferably includes at least 1 of the group consisting of sulfate and chloride, and particularly preferably includes sulfate. By including these salts, the fiber forming ability can be further improved, and the elongation of the resulting fiber can be further improved.

The carboxylate is more preferably sodium carboxylate, the sulfate is more preferably ammonium sulfate, sodium sulfate, magnesium sulfate and calcium sulfate, the chloride is more preferably potassium chloride, sodium chloride, magnesium chloride and calcium chloride, the bicarbonate is more preferably sodium bicarbonate, and the mixed aqueous solution is particularly preferably brackish water and sea water. By using these salts and the mixed aqueous solution, the fiber forming ability can be improved, and the production cost can be further reduced.

The salt content is 0.1 mass% or more, 0.3 mass% or more, 0.5 mass% or more, 0.7 mass% or more, 1 mass% or more, 1.3 mass% or more, 1.5 mass% or more, 1.7 mass% or more, 2 mass% or more, 2.3 mass% or more, 2.5 mass% or more, 2.7 mass% or more, 3 mass% or more, 4 mass% or more, 5 mass% or more, 7 mass% or more, 10 mass% or more, or 15 mass% or more, and the content of the above-mentioned upper limit may be 30 mass% or less, 25 mass% or less, 20 mass% or less, or the solubility or less. For example, the salt content may be 0.1% by mass or more and 30% by mass or less, 0.3% by mass or more and 25% by mass or less, 5% by mass or more and 25% by mass or less, 1% by mass or more and 25% by mass or less, 3% by mass or more and 25% by mass or less, 8% by mass or more and 25% by mass or less, 10% by mass or more and 25% by mass or less, 1% by mass or more and 20% by mass or less, 3% by mass or more and 20% by mass or less, 5% by mass or more and 20% by mass or less, 8% by mass or more and 20% by mass or less, 10% by mass or more and 15% by mass or less, 12% by mass or more and 17% by mass or less, 13% by mass or more and 18% by mass or more, 15% by mass or less and 20% by mass or 16% by mass or more and 20% by mass or less, based on the total amount of the solidification liquid. For example, the salt content is preferably 0.05mol/L or more and 5.5mol/L or less, or 0.1mol/L or more and 5.0mol/L or less, or 0.1mol/L or more and 4.5mol/L or less, or 0.1mol/L or more and 4.0mol/L or less, relative to the total amount of the coagulating liquid.

For example, the salt content in the case of using sodium chloride may be 0.1mol/L or more and 5.0mol/L or less, or 0.1mol/L or more and 4.5mol/L or less, or 0.1mol/L or more and 4.0mol/L or less, relative to the total amount of the coagulating liquid.

For example, the salt content in the case of using sodium sulfate may be 0.1mol/L or more and 3.4mol/L or less, or may be 0.1mol/L or more and 3.0mol/L or less, or may be 0.1mol/L or more and 2.5mol/L or less, or may be 0.1mol/L or more and 2.0mol/L or less, relative to the total amount of the coagulating liquid. For example, the amount of the solidification liquid may be 3% by mass or more and 28% by mass or less, 3% by mass or more and 25% by mass or less, 3% by mass or more and 20% by mass or less, 5% by mass or more and 20% by mass or less, or 8% by mass or more and 20% by mass or less.

The content of sodium sulfate relative to the total amount of the solidification liquid is preferably 10% by mass or more and 20% by mass or less, more preferably 11% by mass or more and 19% by mass or less, still more preferably 11% by mass or more and 18% by mass or less, still more preferably 12% by mass or more and 17% by mass or less, and particularly preferably 13% by mass or more and 16% by mass or less. When the content of sodium sulfate relative to the total amount of the coagulating liquid is 10 mass% or more, a sufficient coagulating speed can be obtained, and an increase in cost due to equipment investment can be avoided. When the content of sodium sulfate relative to the total amount of the coagulating liquid is 20 mass% or less, the occurrence of yarn breakage at the interface between the spinning liquid and the coagulated yarn (yarn) due to rapid coagulation of the spinning liquid can be avoided.

In the above case, the water content corresponding to the total amount of the solidification liquid is preferably 50 mass% or more and 80 mass% or less, more preferably 60 mass% or more and 80 mass% or less, and still more preferably 60 mass% or more and 70 mass% or less, from the viewpoint of improving the recovery efficiency of the solvent. The concentration of the sodium sulfate aqueous solution when sodium sulfate is used is preferably 10% by mass or more and 22% by mass or less, more preferably 10% by mass or more and 20% by mass or less, still more preferably 12% by mass or more and 20% by mass or less, still more preferably 14% by mass or more and 20% by mass or less, and particularly preferably 16% by mass or more and 20% by mass or less. When the concentration of the sodium sulfate aqueous solution is 10 mass% or more, a sufficient solidification speed can be obtained, and an increase in cost due to equipment investment can be avoided. When the concentration of the sodium sulfate aqueous solution is 22 mass% or less, the occurrence of yarn breakage at the interface between the dope and the coagulated yarn (yarn) due to rapid coagulation of the dope can be avoided.

The aqueous solution contained in the coagulation liquid of the present embodiment may be selected from the group consisting of an aqueous carboxylic acid solution, an aqueous bicarbonate solution, an aqueous formate solution, an aqueous acetate solution, an aqueous chloride solution, an aqueous sulfate solution, an aqueous hydrogen phosphate solution, an aqueous citrate solution, brackish water, seawater, and a mixed solution thereof, for example. The aqueous solution contained in the coagulation liquid according to the present embodiment may be selected from the group consisting of, for example, an aqueous citric acid solution, an aqueous formic acid solution, an aqueous sodium bicarbonate solution, an aqueous sodium formate solution, an aqueous sodium acetate solution, an aqueous sodium chloride solution, an aqueous sodium sulfate solution, an aqueous ammonium sulfate solution, an aqueous potassium hydrogen phosphate solution, an aqueous calcium chloride solution, an aqueous sodium citrate solution, brackish water, seawater, and a mixed solution thereof, or may be selected from at least 1 of the group consisting of water, an aqueous formic acid solution, and an aqueous sodium sulfate solution, or may be selected from at least 1 of the group consisting of water and an aqueous sodium sulfate solution.

The coagulation liquid before the contact with the spinning dope may contain an organic solvent or may not contain an organic solvent. When the coagulation liquid contains an organic solvent, the organic solvent may be the same as or different from the organic solvent in the spinning dope, but is preferably the same. In addition, even when the coagulation liquid before the contact with the dope contains no organic solvent, the organic solvent may be dissolved in the coagulation liquid from the contacted dope in the process of bringing the dope into contact with the coagulation liquid. The content of the organic solvent (including the case of dissolving from the dope in the coagulation liquid in contact with the coagulation liquid) contained in the coagulation liquid may be 0 mass% or more and 30 mass% or less, 5 mass% or more and 25 mass% or less, 0 mass% or more and 20 mass% or less, 5 mass% or more and 15 mass% or less, 10 mass% or more and 20 mass% or less, 0 mass% or more and 10 mass% or less, 0 mass% or more and 5 mass% or less, 0 mass% or more and 2 mass% or less, preferably 10 mass% or more and 30 mass% or less, more preferably 12 mass% or more and 28 mass% or more and 14 mass% or less, more preferably 26 mass% or more, based on the total amount of the coagulation liquid (total amount of the organic solvent dissolved from the dope in the coagulation liquid when the dope is dissolved in the coagulation liquid, the total amount of the coagulation liquid is 100 mass% or less. When the content of the organic solvent is within the above range, the fiber forming ability of the structural protein is further improved. As the organic solvent, formic acid, DMSO or HFIP is preferable, formic acid or HFIP is more preferable, and formic acid is further preferable.

The pH of the aqueous solution contained in the coagulating liquid may be 0.25 to 10.00 or 0.25 to 9.50.

The pH of the aqueous acid solution in the coagulating liquid may be, for example, 0.25 to less than 7.00, or 0.50 to less than 7.00, or 1.00 to less than 7.00, or 1.50 to less than 7.00, or 2.00 to less than 7.00, or 3.00 to less than 7.00.

The pH of the brine solution in the coagulating liquid may be, for example, 0.50 to 10.00, 1.00 to 10.00, 2.00 to 10.00, 3.00 to 10.00, 3.50 to 10.00, 4.00 to 10.00, 4.50 to 10.00, 5.00 to 10.00, 5.50 to 10.00, 6.00 to 10.00, 6.50 to 10.00, and 6.50 to 9.50.

The content of the water or aqueous solution in the coagulating liquid may be 60% by mass or more, or 65% by mass or more, or 68% by mass or more, preferably 70% by mass or more, more preferably 71% by mass or more, still more preferably 72% by mass or more, more preferably 73% by mass or more, still more preferably 74% by mass or more, more preferably 75% by mass or more, still more preferably 76% by mass or more, more preferably 77% by mass or more, still more preferably 78% by mass or more, more preferably 79% by mass or more, particularly preferably 80% by mass or more, or 85% by mass or more, or 90% by mass or 95% by mass, based on the total amount of the coagulating liquid. When the content of the above water or aqueous solution is within the above range, the fiber forming ability of the structural protein is further improved. The content of the water or aqueous solution in the solidification liquid may be, for example, 60% by mass or more and 100% by mass or less, or 70% by mass or more and 100% by mass or less, or 75% by mass or more and 100% by mass or less, or 80% by mass or more and 100% by mass or less, or 85% by mass or more and 100% by mass or less, or 90% by mass or more and 100% by mass or less, or 95% by mass or more and 100% by mass or less, or 70% by mass or more and 90% by mass or less, or 75% by mass or more and 85% by mass or less, or 78% by mass or more and 82% by mass or less, based on the total amount of the solidification liquid.

The solidification liquid preferably contains at least 1 of the group consisting of methanol, ethanol, acetone, water and a sulfate aqueous solution, and the content of the methanol, ethanol, acetone, water and/or sulfate aqueous solution in the solidification liquid is preferably 70 mass% or more based on 100 mass% of the total solidification liquid.

The temperature of the solidification liquid may be room temperature or 0 to 90℃or 0 to 80℃or 5 to 80℃or 10 to 80℃or 15 to 80℃or 20 to 80℃or 25 to 80℃or 30 to 80℃or 40 to 80℃or 50 to 80℃or 60 to 80℃or 70 to 80℃or 20 to 70℃or 30 to 70℃or 40 to 70℃or 50 to 70℃or 20 to 60℃or 30 to 60℃or 40 to 60℃or 30 to 50℃or 50 to 60 ℃. When water or an aqueous solution having a PH of 0.25 or more and a PH of 10.00 or less (aqueous formic acid solution, aqueous sodium sulfate solution, or a mixed aqueous solution thereof) is used in the coagulation liquid, the temperature of the coagulation liquid is preferably 30 to 50 ℃, more preferably 32 to 48 ℃, still more preferably 33 to 47 ℃, still more preferably 34 to 46 ℃, still more preferably 35 to 45 ℃ from the viewpoint of more excellent spinning stability. The lower limit of the temperature of the coagulation liquid may be equal to or higher than the melting point of the organic solvent contained in the spinning dope, and the upper limit of the temperature may be equal to or lower than the boiling point of the organic solvent contained in the spinning dope. By increasing the temperature of the coagulation liquid, the desolventizing speed of the spinning dope can be increased.

The coagulation liquid may also contain a dope solvent. The content of the dope solvent (for example, formic acid) relative to the total amount of the coagulation liquid is preferably 15 to 25% by mass, more preferably 16 to 25% by mass, still more preferably 16 to 24% by mass, and particularly preferably 18 to 24% by mass, from the viewpoint of improving the solvent recovery efficiency.

The coagulation liquid may further contain the above-mentioned dissolution promoter which may be added to the spinning dope.

(Method for producing recombinant structural protein multifilament yarn)

(Spinning step)

The method for producing a multifilament according to the present embodiment can be produced by a known wet spinning method, dry-wet spinning method, melt spinning method, or the like using a spinning nozzle having a hole number of 100 or more, and thus a multifilament (a multifilament having a constitution number of 100 or more) composed of the same number of monofilaments as the number of holes of the spinning nozzle can be obtained. The method for producing the multifilament according to the present embodiment can be performed using, for example, a spinning device shown in fig. 6 or 7. The spinning method is preferably wet spinning or dry-wet spinning. The following will describe an example of a method for producing a modified fibroin multifilament yarn.

Fig. 6 is an explanatory view schematically showing an example of a spinning apparatus for producing a modified fibroin multifilament yarn. The spinning device 10 shown in fig. 6 is an example of a spinning device for dry-wet spinning, and includes an extrusion device 1, a coagulation bath 20, a washing bath (drawing bath) 21, and a drying device 4 in this order from the upstream side.

The extrusion device 1 has a reservoir 7 in which a spinning dope (spinning dope) 6 is stored. The coagulation bath 20 stores the coagulation liquid 11. The dope 6 is extruded from a spinning nozzle 9 by a gear pump 8 mounted on the lower end portion of the reservoir 7. The extruded dope 6 is supplied (introduced) into the coagulation liquid 11 of the coagulation bath 20 through the air gap 19. The solvent is removed from the dope in the coagulation liquid 11 to coagulate the modified fibroin and form a fibrous coagulated body. Next, the fibrous coagulated body is supplied to the washing liquid 12 in the washing bath 21, and stretched. The stretch ratio depends on the speed ratio of the first pinch roller 13 and the second pinch roller 14 provided in the washing bath 21. The stretched fibrous coagulated material is then fed into the drying device 4, dried in the yarn path 22, and wound up by a winder. As a result, the multifilament yarn is finally obtained by the spinning device 10 as the wound article 5 wound on the winder. In addition, 18a to 18g are thread guides.

In the case of using a syringe pump having a nozzle with a diameter of 0.1 to 0.6mm as the spinneret 9, the extrusion speed is preferably 0.2 to 6.0 ml/hr per hole, more preferably 1.4 to 4.0 ml/hr. The distance of the coagulated modified fibroin passing through the coagulating liquid 11 (substantially the distance from the wire guide 18a to the wire guide 18 b) may be, for example, 200 to 500mm, as long as the solvent can be removed effectively. The draw-off speed of the undrawn yarn may be, for example, 1 to 100 m/min, or 1 to 20 m/min, and preferably 1 to 3 m/min. When the pull-off speed is 1 m/min or more, productivity can be sufficiently improved. When the extraction speed is 100 m/min or less, significant scattering of the solvent liquid can be avoided. The residence time in the coagulation liquid 11 may be, for example, 0.01 to 3 minutes, and preferably 0.05 to 0.15 minutes, as long as the solvent is removed from the dope. Further, stretching (pre-stretching) may be performed in the solidification liquid 11. The coagulation bath 20 may be provided in a plurality of stages, and stretching may be performed in each stage or in a specific stage as needed.

Fig. 7 is an explanatory view schematically showing an example of a spinning apparatus for producing a modified fibroin multifilament yarn. The spinning device 10 shown in fig. 6 is an example of a spinning device for wet spinning, and is the same as the device of fig. 6 except that the air gap 19 is not provided.

The spinneret shape, the shape of the holes (hole shape), the number of holes (hole number) and the like of the spinning spinneret are not particularly limited, and may be appropriately selected according to the desired fiber diameter, the number of monofilaments and the like.

When the shape of the holes of the spinning nozzle is circular, the pore diameter is exemplified by 0.01mm to 0.6 mm. When the pore diameter is 0.01mm or more, pressure loss can be reduced, and equipment cost can be suppressed. When the pore diameter is 0.6mm or less, the necessity of a stretching operation for reducing the fiber diameter can be reduced, and the possibility of occurrence of stretch breaking during the process from ejection to drawing off can be reduced.

The lower limit value of the number of holes per 1 cone of the spinning nozzle is 100 or more from the viewpoint of improving productivity, preferably 150 or more from the viewpoint of further suitably obtaining the effect, it may be 200 or more, 250 or more, 300 or more, 350 or more, 400 or more, 450 or more, 500 or more, 550 or more, 600 or more, 650 or more, 700 or more, 750 or more, 800 or more, 850 or more, 900 or more, 950 or more, 1000 or more, 1100 or more, 1200 or more, 1300 or more, 1400 or more, 1500 or more, 1600 or more, 1700 or more, 1800 or more, 1900 or more, 400 or more, 2100 or more, 2200 or more, 2300 or more, 2400 or more, 2500 or more, 2600 or more, etc 2700 or more, 2800 or more, 2900 or more, 3000 or more, 3100 or more, 3200 or more, 3300 or more, 3400 or more, 3500 or more, 3600 or more, 3700 or more, 3800 or more, 3900 or more, 4000 or more, 4100 or more, 4200 or more, 4300 or more, 4400 or more, 4500 or more, 4600 or more, 4700 or more, 4800 or more, 4900 or more, 5000 or more, 5100 or more, 5200 or more, 5300 or more, 5400 or more, 5500 or more, 5600 or more, 5700 or more, 5800 or more, 5900 or more.

From the viewpoint of the effect of sufficiently improving productivity, the upper limit value of the number of holes per 1 cone of the spinning nozzle is 9000 or less, or 8900 or less, 8800 or less, 8700 or less, 8600 or less, 8500 or less, 8400 or less, 8300 or less, 8200 or less, 8100 or less, 8000 or less, 7900 or less, 7800 or less, 7700 or less, 7600 or less, 7500 or less, 7400 or less, 7300 or less, 7200 or less, 7100 or less, 7000 or less, 6900 or less, 6800 or less, 6700 or less, 6600 or less, 6500 or less, 6400 or less, 6300 or less, 6200 or less, 6100 or less, 6000 or less.

From the viewpoint of the effect of sufficiently improving productivity, the number of holes per 1 cone of the spinning nozzle may be, for example, 100 or more and 9,000 or less, 150 to 9,000 or less, 200 to 9,000 or less, 250 to 9,000 or less, 350 to 9,000 or less, 300 to 9,000 or less, 350 to 9,000 or less, 400 to 9,000 or less, 450 to 9,000 or less, 500 to 9,000 or less, 650 to 9,000 or less, 750 to 9,000 or less, 800 to 9,000 or less, 850 to 9,000 or less, 900 to 9,000 or less, 950 to 9,000 or 1,000 or less, for example, 100 or more and 8,000 or less, or 150 to 8,000 or less, 200 to 8,000 or less, 250 to 8,000 or less, 350 to 8,000 or less, 300 to 8,000 or less, 350 to 850,000 or less, 350 to 8,000 or less, 800 to 9,000 or less, 850 to 9,000 or 1,000 or less, it may be 150 to 7,000 or less, 200 to 7,000 or less, 250 to 7,000 or less, 350 to 7,000 or less, 300 to 7,000 or less, 350 to 7,000 or less, 400 to 7,000 or less, 450 to 7,000 or less, 500 to 7,000 or less, 650 to 7,000 or less, 750 to 7,000 or less, 800 to 7,000 or less, 850 to 7,000 or less, 900 to 7,000 or less, 950 to 7,000 or 1,000 to 7,000 or less, for example, 100 or more and 6,000 or less, the ratio may be 150 to 6,000 or less, 200 to 6,000 or less, 250 to 6,000 or less, 350 to 6,000 or less, 300 to 6,000 or less, 350 to 6,000 or less, 400 to 6,000 or less, 450 to 6,000 or less, 500 to 6,000 or less, 650 to 6,000 or less, 750 to 6,000 or less, 800 to 6,000 or less, 850 to 6,000 or less, 900 to 6,000 or less, 950 to 6,000 or 1,000 to 6,000 or less, for example, 100 or more and 5,500 or less, or 150 to 5,500 or less, 200 to 5,500 or less, 250 to 5,500 or less, 350 to 5,000 or less, 300 to 5,500 or less, 350 to 5,500 or less, 400 to 5,500 or less, 450 to 5,500 or less, 500 to 5,000 or less, 650 to 5,500 or less, 750 to 5,500 or less, 800 to 5,500 or less, 850 to 5,500 or less, 900 to 5,500 or less, 950 to 5,500 or 1,000 to 5,500 or less, for example, 100 or more and 5,200 or less, 150 to 5,200 or less, 200 to 5,200 or less, 250 to 5,200 or less, 350 to 5,200 or less, 300 to 5,200 or less, 350 to 5,200 or less, 450 to 5,200 or less, 500 to 5,200 or less, 650 to 5,200 or less, 750 to 5,200 or less, 800 to 5,200 or less, 850 to 5,200 or less, 200 to 5,200 or 950 to 5,200 or less, for example, it may be 100 or more and 4,800 or less, or 150 to 4,800 or less, 200 to 4,800 or less, 250 to 4,800 or less, 350 to 4,800 or less, 300 to 4,800 or less, 350 to 4,800 or less, 400 to 4,800 or less, 450 to 4,800 or less, 500 to 4,800 or less, 650 to 4,800 or less, 750 to 4,800 or less, 800 to 4,800 or less, 850 to 4,800 or less, 900 to 4,800 or less, 950 to 4,800 or 1,000 to 4,800 or less, for example, it may be 100 or more and 4,500 or less, or 150 to 4,500 or less, 200 to 4,500 or less, 250 to 4,500 or less, 350 to 4,500 or less, 300 to 4,500 or less, 350 to 4,500 or less, 400 to 4,500 or less, 450 to 4,500 or less, 500 to 4,500 or less, 650 to 4,500 or less, 750 to 4,500 or less, 800 to 4,500 or less, 850 to 4,500 or less, 900 to 4,500 or less, 950 to 4,500 or 1,000 to 4,500 or less, for example, the ratio may be 100 or more and 4,000 or less, or 150 to 4,500 or less, 200 to 4,500 or less, 250 to 4,000 or less, 350 to 4,500 or less, 300 to 4,000 or less, 350 to 4,000 or less, 400 to 4,000 or less, 450 to 4,000 or less, 500 to 4,000 or less, 650 to 4,000 or less, 750 to 4,000 or less, 800 to 4,000 or less, 850 to 4,500 or less, 900 to 4,000 or less, and, 950 to 4,000 or less or 1,000 to 4,000 or less.

The number of spinning nozzles may be appropriately selected depending on the amount of the target fiber, and is not particularly limited, and may be, for example, 1,2, 3, 4,5, 6,7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30, 35, 40, 45, 60, or more, 95, or more, or 80, or more, and the number of spinning nozzles may be appropriately selected. The spinning nozzle may be a combination of a plurality of spinning nozzles (a plurality of spindles) having the same number of holes (for example, a combination of 12 spinning nozzles having a number of holes of 3,000), or a combination of a plurality of spinning nozzles having different numbers of holes.

The temperature of the spinning dope and the temperature of the spinning nozzle when passing through the spinning nozzle are not particularly limited, and can be appropriately adjusted according to the concentration and viscosity of the spinning dope to be used, the kind of the organic solvent, and the like. From the viewpoint of preventing degradation of modified fibroin, the temperature is preferably 30 to 100 ℃. In addition, from the viewpoint of reducing the possibility of pressure rise due to volatilization of the solvent and clogging in the piping due to solidification of the dope, the upper limit value of the temperature is preferably smaller than the boiling point of the solvent used. This improves the process stability.

The temperature of the solidification liquid 11 is not particularly limited, and may be 40℃or lower, 30℃or lower, 25℃or lower, 20℃or lower, 10℃or lower, or 5℃or lower. From the viewpoints of operability, cooling cost, and the like, it is preferably 0 ℃ or higher. The temperature of the coagulation liquid 11 can be adjusted by using, for example, a spinning device 10 having a coagulation bath 20 having a heat exchanger therein and a cooling circulation device. For example, by passing the medium cooled to a predetermined temperature by the cooling circulation device through the heat exchanger provided in the coagulation bath, the temperature can be adjusted to be within the above-described range by heat exchange between the coagulation liquid 11 and the heat exchanger. In this case, by circulating the solvent which is the medium and is used in the solidification liquid 11, cooling can be performed more effectively.

A plurality of coagulation baths for storing the coagulation liquid may be provided.

The modified fibroin (fibrous coagulated body) after coagulation may be taken up directly by a winder after being separated from the coagulation bath or the washing bath, or may be taken up by a winder after being dried by a drying device.

The modified fibroin (fibrous coagulated body) after coagulation may be determined by the extrusion speed (ejection speed) of the spinning dope from the nozzle, and the like, as long as the solvent can be efficiently removed by the distance in the coagulating liquid. The residence time of the coagulated modified fibroin (or spin dope) in the coagulation liquid may be determined according to the distance of the coagulated modified fibroin in the coagulation liquid, the extrusion speed of the spin dope from the nozzle, and the like.

(Stretching step)

The method for producing the modified fibroin multifilament according to the present embodiment may further include a step (stretching step) of stretching and solidifying the modified fibroin (fibrous solidified body). The stretching method includes wet heat stretching, dry heat stretching, and the like. The stretching step may be performed in, for example, the coagulation bath 20 or the washing bath 21. The stretching step may be performed in air.

The stretching performed in the washing bath 21 may be so-called wet-heat stretching performed in warm water, a solution in which an organic solvent or the like is added to warm water, or the like. The temperature of the wet heat stretching is preferably 50 to 90 ℃. When the temperature is 50 ℃ or higher, the pore diameter of the yarn can be slightly stabilized. In addition, when the temperature is 90 ℃ or less, the temperature can be easily set, and the stability of spinning can be improved. The temperature is more preferably 75 to 85 ℃.

The wet heat stretching may be performed in warm water, a solution in which an organic solvent or the like is added to warm water, or steam heating. The temperature may be, for example, 40 to 200 ℃, 50 to 180 ℃, 50 to 150 ℃, or 75 to 90 ℃. The draw ratio at the time of wet-heat drawing may be, for example, 1 to 30 times, or 2 to 25 times, or 2 to 20 times, or 2 to 15 times, or 2 to 10 times, or 2 to 8 times, or 2 to 6 times, or 2 to 4 times, relative to the undrawn yarn (or pre-drawn yarn). However, the draw ratio is not limited as long as it is within a range that can obtain desired properties such as the thickness and mechanical properties of the fiber.

The dry heat stretching may be performed by stretching in air using a device provided with a heat source such as a contact hot plate or a non-contact furnace, but is not particularly limited as long as the stretching is performed at a predetermined magnification while heating the fiber to a predetermined temperature. The temperature may be, for example, 100℃to 270℃or 140℃to 230℃or 140℃to 200℃or 160℃to 180 ℃.

The draw ratio in the dry heat drawing step may be, for example, 1 to 30 times, 2 to 20 times, or 3 to 15 times, preferably 3 to 10 times, more preferably 3 to 8 times, and even more preferably 4 to 8 times, relative to the undrawn yarn (or pre-drawn yarn). However, the draw ratio is not limited as long as it is within a range that can obtain desired properties such as the thickness and mechanical properties of the fiber.

In the stretching step, wet heat stretching and dry heat stretching may be performed separately, or may be performed in a plurality of stages, or may be performed in combination. That is, as the stretching step, the first-stage stretching may be performed by wet-heat stretching, the second-stage stretching may be performed by dry-heat stretching, or the first-stage stretching may be performed by wet-heat stretching, the second-stage stretching may be performed by wet-heat stretching, the third-stage stretching may be performed by dry-heat stretching, and the wet-heat stretching and the dry-heat stretching may be appropriately combined.

The lower limit value of the final draw ratio of the multifilament after the drawing step is preferably 1,2,3, 4, 5, 6, 7, 8 or 9 times as compared to the undrawn yarn (or the pre-drawn yarn). The upper limit value of the final draw ratio of the multifilament after the drawing step is preferably 40 times, 30 times, 20 times, 15 times, 14 times, 13 times, 12 times, 11 times or 10 times. For example, the final stretch ratio may be 3 to 40 times, 3 to 30 times, 5 to 20 times, 5 to 15 times, or 5 to 13 times. However, the draw ratio is not limited as long as it is within a range that can obtain desired properties such as the thickness and mechanical properties of the fiber. By adjusting the draw ratio, the fiber diameter of the obtained multifilament can be adjusted to an arbitrary value.

Before or after drying, an oiling agent may be applied to the undrawn yarn (or the pre-drawn yarn) or the drawn yarn for the purpose of imparting charge-suppressing property, astringency, lubricity, and the like, as required. The type of the oil to be added and the amount to be added are not particularly limited, and may be appropriately adjusted in consideration of the use of the fiber, the operability of the fiber, and the like.

The production method according to the present embodiment may further include a step of filtering the dope before the discharge of the dope (filtering step), and/or a step of defoaming the dope before the discharge (defoaming step).

The manufacturing method according to the present embodiment may further include a shrinking step before or after the drying step. In the case of performing the shrinking step after the drying step, the spun dry multifilament yarn may be wound around a bobbin, and then the spun multifilament yarn may be wound out of the bobbin to perform the shrinking step.

(Shrinkage step)

The modified fibroin multifilament according to the present embodiment may further include a shrinking step of irreversibly shrinking the multifilament. In the shrinkage step of irreversibly shrinking the multifilament yarn, the multifilament yarn may be irreversibly shrunk by bringing the multifilament yarn into contact with water, and/or the multifilament yarn may be irreversibly shrunk by heat-relaxing the multifilament yarn. In the case where the multifilament yarn is irreversibly contracted by contact with water, the multifilament yarn after the irreversible contraction is dried and further contracted.

(Shrinking step (contacting step) with Water)

Fig. 8 is a diagram showing an example of a change in the length of a multifilament yarn (including a fiber modified with fibroin) caused by contact with water. The multifilament yarn (modified fibroin-containing fiber) according to the present embodiment has a characteristic of shrinking (primary shrinkage) by contact with water having a boiling point lower than that (wetting) (in fig. 8, the length change indicated by "primary shrinkage"). When drying is performed after the primary shrinkage, further shrinkage (in fig. 8, the length change indicated by "secondary shrinkage") is performed. When the film is contacted with water again after the secondary shrinkage, the film is elongated to a length equal to or similar to that before the secondary shrinkage, and when the drying and wetting are repeated thereafter, the shrinkage and elongation are repeated at a width (width indicated by "expansion and contraction rate" in fig. 8) equal to that of the secondary shrinkage. That is, the primary shrinkage caused by contacting the multifilament yarn with water is an irreversible shrinkage. Therefore, in the shrinking step, the multifilament is brought into contact with water, whereby a modified fibroin multifilament having a shrinking history in which irreversible shrinkage occurs according to the present embodiment can be obtained. The step of bringing the multifilament into contact with water to cause irreversible shrinkage (primary shrinkage) is hereinafter referred to as a "contact step".

The irreversible shrinkage of the multifilament yarn (including the modified fibroin-containing fibers) in the contacting step (the "primary shrinkage" in fig. 8) is considered to occur, for example, for the following reasons. That is, one of the reasons is considered to be that the multifilament (fiber containing modified fibroin) is caused by a primary structure of the multifilament (fiber containing modified fibroin), and the other is considered to be that the multifilament (fiber containing modified fibroin) having residual stress by, for example, stretching in a production process is immersed in water between fibers or in fibers, thereby relaxing the residual stress.

In the contacting step, after spinning, the multifilament before being contacted with water is contacted with water so that the multifilament is in a wet state. The wet state means a state in which at least a part of the multifilament is wetted with water. This makes it possible to shrink the multifilament yarn without depending on an external force. The shrinkage is irreversible (corresponding to the "primary shrinkage" of fig. 8).

In the contacting step, the temperature of the water in contact with the multifilament yarn may be lower than the boiling point. Thereby, workability in a shrinkage process, and the like are improved. In view of sufficiently shortening the shrinkage time, the lower limit of the temperature of water is preferably 10 ℃ or higher, more preferably 40 ℃ or higher, and still more preferably 70 ℃ or higher. The upper limit of the temperature of water is preferably 90 ℃ or lower.

In the contacting step, the method of contacting the multifilament with water is not particularly limited. Examples of the method include a method of immersing the multifilament in water, a method of spraying the multifilament with water at normal temperature or in a state of heating steam, and a method of exposing the multifilament to a high humidity atmosphere filled with water vapor. Among these methods, in the contacting step, a method of immersing the multifilament in water is preferable, because shortening of the shrinking time can be effectively achieved and simplification of processing equipment and the like can be achieved.

In the contacting step, when the multifilament is contacted with water in a relaxed state, the multifilament is not only shrunk but also waved and wrinkled. In order to prevent such wrinkling, for example, the multifilament is stretched in the fiber axis direction without applying tension to be brought into contact with water, or the like, the contacting step may be performed without loosening the multifilament.

(Drying step)

The method for producing the modified fibroin multifilament according to the present embodiment may be a method including a drying step. The drying step is a step of drying and further shrinking the multifilament yarn subjected to the contacting step (or the modified fibroin multifilament yarn obtained by the contacting step) (corresponding to "secondary shrinkage" in fig. 8). The drying may be, for example, natural drying or forced drying using a drying device. As the drying apparatus, any known contact type or non-contact type drying apparatus may be used. The drying temperature is not limited as long as it is lower than the temperature at which the modified fibroin contained in the multifilament is decomposed or the multifilament is thermally damaged, and is generally in the range of 20 to 150 ℃, preferably 50 to 100 ℃. By having the temperature within this range, thermal damage to the fibers or decomposition of the modified fibroin contained in the fibers does not occur, and the fibers can be dried more rapidly and effectively. The drying time is appropriately set in accordance with the drying temperature and the like, and for example, a time period or the like is employed in which the influence on the quality, physical properties, and the like of the modified fibroin multifilament due to excessive drying can be reduced as rapidly as possible.

Fig. 9 is an explanatory view schematically showing an example of a production apparatus for producing the modified fibroin multifilament yarn. The manufacturing apparatus 40 shown in fig. 9 includes a feed roller 42 for feeding the multifilament, a winder 44 for winding the modified fibroin multifilament 38, a water bath 46 for performing the contacting step, and a dryer 48 for performing the drying step.

More specifically, the feed roller 42 may be provided with a wound product of the multifilament 36, and the multifilament 36 may be continuously and automatically fed from the wound product of the multifilament 36 by rotation of a motor or the like, not shown. The winder 44 continuously and automatically winds the modified fibroin multifilament 38 produced by the contact process and the drying process after being fed out from the feed roller 42 by the rotation of a motor not shown. In addition, the feed speed at which the multifilament yarn 36 is fed by the feed roller 42 and the winding speed at which the modified fibroin multifilament yarn 38 is wound by the winder 44 can be controlled independently of each other.

The water bath 46 and the dryer 48 are disposed in parallel on the upstream side and downstream side in the feeding direction of the multifilament yarn 36 between the feed roller 42 and the winder 44, respectively. The manufacturing apparatus 40 shown in fig. 9 includes transfer rollers 50 and 52 for transferring the multifilament 36 before and after the contact step of traveling from the feed roller 42 to the winder 44.

The water bath 46 has a heater 54, and the water 47 heated by the heater 54 is contained in the water bath 46. Further, in the water bath 46, a tension roller 56 is provided in a state immersed in the water 47. Thus, the multifilament 36 fed from the feed roller 42 is immersed in the water 47 in a state wound around the tension roller 56 in the water bath 46, and travels toward the winder 44. In addition, the immersion time of the multifilament yarn 36 in the water 47 is appropriately controlled according to the traveling speed of the multifilament yarn 36.

Dryer 48 has a pair of heated rolls 58. A pair of heated rollers 58 can wind the multifilament yarn 36 which is released from the water bath 46 and travels toward the winder 44. Thus, the multifilament 36 immersed in the water 47 in the water bath 46 is heated in the dryer 48 by the pair of hot rolls 58, dried, and then sent to the winder 44.

When the modified fibroin multifilament 38 is manufactured using the manufacturing apparatus 40 having such a structure, first, for example, a wound product of the multifilament 36 spun using the spinning apparatus 10 shown in fig. 11 is mounted on the feed roller 42. Multifilament yarn 36 is then continuously fed from feed roll 42 and immersed in water 47 in a water bath 46. At this time, for example, the winding speed of the winder 44 is made slower than the feeding speed of the feed roller 42. Thus, since the multifilament 36 is contracted by contact with the water 47 in a state of not being loosened between the feed roller 42 and the winder 44, occurrence of wrinkling can be prevented. Upon contact with water 47, the multifilament yarn 36 irreversibly contracts (corresponding to "primary contraction" of fig. 8).

Next, the multifilament 36 after being contacted with the water 47 (or the modified fibroin multifilament 38 manufactured through the contact with the water 47) is heated by a pair of hot rollers 58 of the dryer 48. This can dry the multifilament 36 after the contact with water 47 (or the modified fibroin multifilament 38 produced by the contact with water 47) and shrink it (corresponding to "secondary shrink" in fig. 8). At this time, the ratio of the feeding speed of the feed roller 42 to the winding speed of the winder 44 may be controlled so that the length of the modified fibroin multifilament 38 does not change. The obtained modified fibroin multifilament 38 is then wound with a winder 44 to obtain a wound product of the modified fibroin multifilament 38.

Instead of the pair of heat rollers 58, a drying device such as a dry heat plate 64 shown in fig. 10 (b) which is constituted only by a heat source may be used to dry the multifilament 36 after being contacted with the water 47. In this case, as in the case of using the pair of heating rollers 58 as the drying means, the length of the modified fibroin multifilament yarn can be prevented from being changed by adjusting the relative speed between the feeding speed of the feed roller 42 and the winding speed of the winder 44. The drying unit is formed by a drying plate 64. In addition, a dryer 48 is not necessary.

As described above, by using the manufacturing apparatus 40, the intended modified fibroin multifilament 38 can be automatically, continuously, and extremely easily manufactured.

Fig. 10 is an explanatory view schematically showing another example of a production apparatus for producing the modified fibroin multifilament yarn. Fig. 10 (a) shows a processing apparatus for performing a contact process (primary shrinkage) provided in the manufacturing apparatus, and fig. 10 (b) shows a drying apparatus for performing a drying process provided in the manufacturing apparatus. The manufacturing apparatus shown in fig. 10 has a processing apparatus 60 for performing a contact process on the multifilament yarn 36 and a drying apparatus 62 for drying the multifilament yarn 36 after the contact process (or the modified fibroin multifilament yarn 38 manufactured through the contact process), and has a structure independent of each other.

More specifically, the processing device 60 shown in fig. 10 (a) has a structure in which the feed roller 42, the water bath 46, and the winder 44 are arranged in this order from the upstream side to the downstream side in the traveling direction of the multifilament yarn 36. This processing device 60 dips the multifilament yarn 36 fed from the feed roll 42 into the water 47 in the water bath 46 to cause it to shrink. Then, the modified fibroin multifilament 38 thus obtained was wound up by a winder 44. At this time, for example, the winding speed of the winder 44 is made slower than the feeding speed of the feed roller 42. Accordingly, the multifilament 36 is in contact with the water 47 in a relaxed state between the feed roller 42 and the winder 44, and thus is contracted, whereby the application of tension to the fibers can be prevented. Upon contact with water 47, the multifilament yarn 36 irreversibly contracts (corresponding to "primary contraction" of fig. 8).

The drying apparatus 62 shown in fig. 10 (b) has a feed roller 42, a winder 44, and a drying plate 64. The dry heat plate 64 is disposed between the feed roller 42 and the winder 44 such that the dry heat surface 66 is in contact with the modified fibroin multifilament yarn 38 and extends along the traveling direction thereof. In the drying device 62, as described above, the length of the modified fibroin multifilament 38 may not be changed by controlling the ratio of the feed-out speed of the feed roller 42 and the winding speed of the winder 44, for example.

By using the manufacturing apparatus having such a structure, after the multifilament 36 is contracted by the processing apparatus 60 to obtain the modified fibroin multifilament 38, the modified fibroin multifilament 38 can be dried by the drying apparatus 62.

The feeding roller 42 and the winding machine 44 may be omitted from the processing device 60 shown in fig. 10 (a), and the processing device may be composed of only the water bath 46. In the case of using a manufacturing apparatus having such a processing apparatus, for example, a modified fibroin multifilament yarn is manufactured by a so-called batch type. Further, the drying device 62 shown in fig. 10 (b) is not essential.

(Shrinkage step due to thermal relaxation)

The shrinkage step of irreversibly shrinking the multifilament yarn may be performed by heating and relaxing the multifilament yarn. The thermal relaxation of the multifilament can be performed by heating the multifilament to relax the multifilament in a heated state and then shrinking the multifilament. In the following, the step of heating the multifilament in the shrinkage due to thermal relaxation of the multifilament is referred to as a "heating step", and the step of relaxing the multifilament in a heated state and then shrinking is referred to as a "relaxation shrinkage step". The heating step and the relaxation and contraction step can be performed by, for example, heating the relaxation device 140 at a high temperature as shown in fig. 11 and 12.

(Heating step)

In the heating step, the heating temperature of the multifilament yarn 36 is preferably equal to or higher than the softening temperature of the modified fibroin used for the multifilament yarn 36. The softening temperature of the modified fibroin in the present specification means a temperature at which shrinkage starts due to stress relaxation of the multifilament yarn 36. In the heat relaxation shrinkage of the modified fibroin at a softening temperature or higher, the fiber is shrunk to such an extent that the fiber cannot be obtained by merely releasing the moisture in the fiber, whereby the residual stress in the fiber due to the stretching in the spinning process can be removed.

The temperature corresponding to the softening temperature may be 180 ℃. When the thermal relaxation shrinkage is performed in a high temperature range of 180 ℃ or more, the larger the relaxation rate or the higher the temperature, the more effective the residual stress in the multifilament can be removed. Therefore, the heating temperature of the multifilament yarn 36 is preferably 180℃or higher, more preferably 180℃to 280℃and still more preferably 200℃to 240℃and particularly preferably 220℃to 240 ℃.

The heating time in the heating step, that is, the residence time in the high-temperature heating furnace 143 is preferably 60 seconds or less, more preferably 30 seconds or less, and even more preferably 5 seconds or less, from the viewpoint of not impairing the elongation of the fiber after the heat treatment. The length of this heating time is considered to have little effect on stress. In addition, when the heating temperature is 200 ℃ and the heating time is 5 seconds or less, the fiber elongation after the heat treatment can be prevented from decreasing.

(Relaxation shrinkage Process)

In the relaxation shrinkage step, the relaxation magnification is preferably more than 1 time, more preferably 1.4 times or more, further preferably 1.7 times or more, and particularly preferably 2 times or more. The relaxation rate is a ratio of the feed-out speed to the winding speed of the multifilament yarn 36, and more specifically, a ratio of the feed-out speed of the feed-out roller 141 to the winding speed of the winding roller 142.

In the thermal relaxation method using the high-temperature thermal relaxation device 140, the thermal relaxation step and the relaxation shrinkage step may be performed separately as long as the thermal relaxation can be performed while heating the multifilament 36. That is, the heating device may be a separate and independent device from the relaxing device. In this case, a relaxation device is provided at the rear stage (downstream side in the traveling direction of the multifilament yarn 36) of the heating device in order to perform the relaxation and contraction process after the heating process.

The thermal relaxation step may be performed on the multifilament separately from the production step of the multifilament. That is, as a separate device provided separately from the spinning device 25, the same device as the high-temperature heating and relaxing device 140 may be provided. It is also possible to use a method in which the multifilament yarn 36 manufactured separately is set on and fed out from a feeding roller. The heat relaxing step may be performed on 1 multifilament or may be performed on a plurality of bundled filaments.

(Crosslinking step)

The crosslinking process for chemical crosslinking between polypeptide molecules in the fiber may be further performed for the modified fibroin multifilament having a shrinkage history of irreversible shrinkage obtained as described above, or for the multifilament before irreversible shrinkage. Examples of the functional group capable of crosslinking include amino groups, carboxyl groups, thiol groups, and hydroxyl groups. For example, the amino group of a lysine side chain contained in a polypeptide may be crosslinked with the carboxyl group of a glutamic acid or aspartic acid side chain by dehydration condensation as an amide bond. The crosslinking may be performed by a dehydration condensation reaction under vacuum heating, or may be performed by a dehydration condensing agent such as carbodiimide.

The crosslinking between polypeptide molecules may be performed using a crosslinking agent such as carbodiimide or glutaraldehyde, or may be performed using an enzyme such as transglutaminase. The carbodiimide is a compound represented by the general formula R ₁N＝C＝NR₂ (wherein R ₁ and R ₂ each independently represent an organic group including an alkyl group or a cycloalkyl group having 1 to 6 carbon atoms). Specific examples of the carbodiimide include 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), N' -Dicyclohexylcarbodiimide (DCC), 1-cyclohexyl-3- (2-N-morpholinoethyl) carbodiimide, and Diisopropylcarbodiimide (DIC). Among these, EDC and DIC are preferable because they have high amide bond forming ability between polypeptide molecules and are easy to undergo crosslinking reaction.

The crosslinking treatment preferably imparts a crosslinking agent to the fibers and crosslinks the fibers under vacuum heat drying. The crosslinking agent may be added to the fiber as a pure product or may be diluted to a concentration of 0.005 to 10 mass% with a lower alcohol having 1 to 5 carbon atoms, a buffer, or the like. The crosslinking treatment is preferably carried out at a temperature of 20 to 45℃for 3 to 42 hours. By the crosslinking treatment, a higher stress (strength) can be imparted to the fiber.

(Multifilament yarn)

The multifilament according to the present embodiment contains a recombinant structural protein, has 100 or more constituent filaments, and has a coefficient of variation in elongation of less than 33%. The constituent filaments herein refer to filaments constituting the multifilament (also referred to as filaments), and the constituent number refers to the number of filaments constituting the multifilament (the number of filaments). The multifilament according to the present embodiment contains, for example, modified fibroin, and the number of the multifilament is 100 or more, the coefficient of variation of the elastic modulus of the multifilament may be 15% or less, the coefficient of variation of the strength of the multifilament may be 15% or less, and the coefficient of variation of the elongation of the multifilament is less than 33%.

From the viewpoint of improving productivity, the lower limit of the number of constituent multifilament yarns is 100 or more. From the viewpoint of further suitably obtaining the effect, it is preferably 150 or more, it may be 200 or more, 250 or more, 300 or more, 350 or more, 400 or more, 450 or more, 500 or more, 550 or more, 600 or more, 650 or more, 700 or more, 750 or more, 800 or more, 850 or more, 900 or more, 950 or more, 1,000 or more, 1,100 or more, 1,200 or more, 1,300 or more, 1,400 or more, 1,500 or more, 1,600 or more, 1,700 or more, 1,800 or more, 1900 or more, 2,000 or more, 2,100 or more, 2,200 or more, 2,300 or more, 2,400 or more, 2,500 or more, 2,600 or more, or more 2700 or more, 2,800 or more, 2,900 or more, 3000 or more, 3100 or more, 3,200 or more, 3300 or more, 3400 or more, 3,500 or more, 3,600 or more, 3,700 or more, 3,800 or more, 3900 or more, 4000 or more, 4,100 or more, 4200 or more, 4,300 or more, 4,400 or more, 4500 or more, 4600 or more, 4,700 or more, 4,800 or more, 4,900 or more, 5,000 or more, 5,100 or more, 5,200 or more, 5,300 or more, 5,400 or more, 5,500 or more, 5,600 or more, 5,700 or more, 5,800 or more, 5,900 or more, the number of holes of the spinning nozzle and/or the number of spindles of the spinning nozzle may be appropriately selected according to the use.

From the viewpoint of the effect of sufficiently improving productivity, the upper limit value of the number of constituent multifilament yarns is 900,000 or less, the values may be 800,000 or less, 700,000 or less, 675,000 or less, 600,000 or less, 500,000 or less, 450,000 or less, 400,000 or less, 300,000 or less, 250,000 or less, 200,000 or less, 150,000 or less, 100,000 or less, 98,000 or less, 96,000 or less, 95,000 or less, 90,000 or less, 85,000 or less, 80,000 or less, 78,000 or less, 76,000 or less, 75,000 or less, 72,000 or less, 70,000 or less, 68,000 or less, 66,000 or less, 64,000 or less, 62,000 or less, 60,000 or less, 58,000 or less, 56,000 or less, 54,000 or less, 52,000 or less, 50,000 or less, 48,000 or less, 46,000 or less, 45,000 or less, 40,000 or less, 38,000 or less, 36,000 or less. 32,000 or less, 30,000 or less, 28,000 or less, 26,000 or less, 25,000 or less, 24,000 or less, 22,000 or less, 20,000 or less, 18,000 or less, 16,000 or less, 14,000 or less, 12,000 or less, 10,000 or less, 9,500 or less, 9,000 or less, 8,900 or less, 8,800 or less, 8,700 or less, 8600 or less, 8500 or less, 8400 or less, 8,300 or less, 8,200 or less, 8,100 or less, 8,000 or less, 7,900 or less, 7800 or less, 7,700 or less, 7,600 or less, 7500 or less, 7,400 or less, 7300 or less, 7,200 or less, 7,100 or less, 7,000 or less, 6,900 or less, 6,800 or less, 6,700 or less, 6,600 or less, 6,500 or less.

From the viewpoint of the effect of sufficiently improving productivity, the number of constituent multifilament is 100 to 900,000 or less, and for example, 100 to 800,000, 100 to 700,000, 100 to 600,000, 100 to 500,000, 100 to 400,000, 100 to 300,000, 100 to 200,000, 100 to 100,000, 1,000 to 90,000, 950 to 90,000, 900 to 90,000, 800 to 90,000, 700 to 90,000, 600 to 90,000, 550 to 90,000, 500 to 90,000, 450 to 90,000, 400 to 90,000, 350 to 90,000, 300 to 90,000, 250 to 90,000, 200 to 90,000, 150 to 90,000 or 100 to 90,000, but also 1,000 to 60,000, 950 to 60,000, 900 to 60,000, 850 to 60,000, 800 to 60,000, 750 to 60,000, 700 to 60,000, 650 to 60,000, 600 to 60,000, 550 to 60,000, 500 to 60,000, 450 to 60,000, 400 to 60,000, 350 to 60,000, 300 to 60,000, 250 to 60,000, 200 to 60,000, 150 to 60,000 or 100 to 60,000, but also 1,000 to 48,000, 950 to 48,000, 900 to 48,000, 850 to 48,000, 800 to 48,000, 750 to 48,000, 700 to 48,000, 650 to 48,000, 600 to 48,000, 550 to 48,000, 500 to 48,000, 450 to 48,000, 400 to 48,000, 350 to 48,000, 300 to 48,000, 250 to 48,000, 200 to 48,000, 150 to 48,000 or 100 to 48,000, it may be 1,000 to 39,000, 950 to 39,000, 900 to 39,000, 850 to 39,000, 800 to 39,000, 750 to 39,000, 700 to 39,000, 650 to 39,000, 600 to 39,000, 550 to 39,000, 500 to 39,000, 450 to 39,000, 400 to 39,000, 350 to 39,000, 300 to 39,000, 250 to 39,000, 200 to 39,000, 150 to 39,000 or 100 to 39,000, 1,000 to 30,000, 950 to 30,000, 900 to 30,000, 850 to 30,000, 800 to 30,000, 750 to 30,000, 700 to 30,000, 650 to 30,000, 600 to 30,000, 550 to 30,000, 500 to 30,000, 450 to 30,000, 400 to 30,000, 350 to 30,000, 300 to 30,000, 250 to 30,000, 200 to 30,000, 150 to 30,000, or 100 to 30,000, but may also be 1,000 to 24,000, 950 to 24,000, 900 to 24,000, 850 to 24,000, 800 to 24,000, 750 to 24,000, 700 to 24,000, 650 to 24,000, 600 to 24,000, 550 to 24,000, 500 to 24,000, 450 to 24,000, 400 to 24,000, 350 to 24,000, 300 to 24,000, 250 to 24,000, 200 to 24,000, 150 to 24,000, or 100 to 24,000, but also 1,000 to 18,000, 950 to 18,000, 900 to 18,000, 850 to 18,000, 800 to 18,000, 750 to 18,000, 700 to 18,000, 650 to 18,000, 600 to 18,000, 550 to 18,000, 500 to 18,000, 450 to 18,000, 400 to 18,000, 350 to 18,000, 300 to 18,000, 250 to 18,000, 200 to 18,000, 150 to 18,000 or 100 to 18,000, but also 1,000 to 12,000, 950 to 12,000, 900 to 12,000, 850 to 12,000, 800 to 12,000, 750 to 13,000, 700 to 12,000, 650 to 12,000, 600 to 12,000, 550 to 12,000, 500 to 12,000, 450 to 12,000, 400 to 12,000, 350 to 12,000, 300 to 12,000, 250 to 12,000, 200 to 12,000, 150 to 12,000 or 100 to 12,000, 1,000 ～ 100,000, 950 to 10,000, 900 to 10,000, 850 to 10,000, 800 to 10,000, 750 to 10,000, 700 to 10,000, 650 to 10,000, 600 to 10,000, 550 to 10,000, 500 to 10,000, 450 to 10,000, 400 to 10,000, 350 to 10,000, 300 to 10,000, 250 to 10,000, 200 to 10,000, 150 to 10,000 or 100 to 10,000, 1,000 to 9,000, 950 to 9,000, 900 to 9,000, 850 to 9,000, 800 to 9,000, 750 to 9,000, 700 to 9,000, 650 to 9,000, 600 to 9,000, 550 to 9,000, 500 to 4,000, 450 to 9,000, 400 to 9,000, 350 to 9,000, 300 to 9,000, 250 to 9,000, 200 to 9,000, 150 to 9,000 or 100 to 9,000, but may be 1,000 to 6,000, 950 to 6,000, 900 to 6,000, 850 to 6,000, 800 to 6,000, 750 to 6,000, 700 to 6,000, 650 to 6,000, 600 to 6,000, 550 to 6,000, 500 to 6,000, 450 to 6,000, 400 to 6,000, 350 to 6,000, 300 to 6,000, 250 to 6,000, 200 to 6,000, 150 to 6,000 or 100 to 6,000.

The coefficient of variation of the elastic modulus, the coefficient of variation of the strength, the coefficient of variation of the elongation, and the coefficient of variation of the fineness of the multifilament can be calculated by measuring the elastic modulus [ gf/D ] ([ gf/den ]), the strength [ g/D ] ([ g/den ]), the elongation at break [% ], and the fineness [ D ] ([ den ]), to determine the average value and the standard deviation of each physical property, respectively. The calculation of each coefficient of variation is performed using the following equation. The smaller the value of each coefficient of variation means the smaller the variation of each physical property value.

Coefficient of variation of elastic modulus (CV) [% ] = standard deviation of elastic modulus/average value of elastic modulus x 100

Coefficient of variation of intensity (CV) [% ] = standard deviation of intensity/average value of intensity x 100

Coefficient of variation of elongation (CV) [% ] = standard deviation of elongation/average value of elongation x 100

Coefficient of variation of titer (CV) [% ] = standard deviation of titer/mean of titer x 100

The elastic modulus [ gf/D ], the strength [ g/D ], and the elongation at break [% ] of the multifilament and the calculation of the respective standard deviations can be performed according to JIS L1013 using a 3345 series tensile tester manufactured by Instron corporation. For example, the test may be performed under an environment having a temperature of 20℃and a relative humidity of 65% and a test length of 300mm and a test speed of 300 mm/min as test conditions, and the capacity of the load cell may be appropriately selected according to the fineness of the multifilament (fiber). The measurement value may be calculated as an average value of, for example, the sampling number n=5.

The upper limit value of the coefficient of variation of the elongation of the multifilament is less than 33%. When the coefficient of variation in elongation is less than 33%, the process passability in the post-process steps such as twisting, spinning, weaving, braiding, or cutting can be further improved. The lower limit value of the coefficient of variation in elongation of the multifilament may be 0.01% or more, 0.1% or more, 0.2% or more, 0.3% or more, 0.4% or more, 0.5% or more, 0.6% or more, 0.7% or more, 0.8% or more, 0.9% or more, 1% or more, 1.3% or more, 1.4% or more, 1.5% or more, 1.6% or more, 1.7% or more, 1.8% or more, 1.9% or more, 2% or more, 2.1% or more, 2.2% or more, 2.3% or more, 2.4% or more, or 2.5% or more, and may be appropriately selected in view of productivity. The coefficient of variation in elongation of the multifilament is preferably 32% or less, 31% or less, 30% or less, 29% or less, 28% or less, 27% or less, 26% or less, 25% or less, 24% or less, or 23% or less, more preferably 22% or less, 21% or less, 20% or less, 19% or less, 18% or less, 17% or less, 16% or less, 15% or less, 14% or less, 13% or less, 12% or 11% or less, and still more preferably 10% or less, 9.5% or less, 9% or less, 8.5% or less, 8% or less, 7.5% or less, 7% or less, 6.5% or less, 6% or less, 5.5% or less.

Further, the elongation coefficient of the multifilament yarn is preferably 0.01% to less than 0.1%, more than 0.1% and less than 1%, more than 1% and less than 5%, more than 5% and less than 10%, more than 10% and less than 15%, more preferably 0.01% to less than 0.1%, more than 0.1% and less than 1%, more preferably 1% and less than 5%, more preferably 5% and less than 10%, more than 10% and less than 15%, more preferably 15% and less than 20% or more than 20% and less than 25%, even more preferably 0.01% to less than 0.1%, more preferably 0.1% and less than 10%, more than 10% and 15% or more than 15%, more preferably 0.01% to less than 0.1%, more preferably 0.1% and less than 1%, more than 1% and more preferably 0.1% and more than 10%, more than 1% and more preferably 0.1% and less than 1%, more than 1% and more preferably 0.1% and less than 15% and more than 20% and less than 25%, still more preferably 0.01% to less than 0.1%, more than 10% and less than 10% and more than 15% and more preferably 0.1%.

The elongation coefficient of the multifilament is preferably 0.01 to less than 33％、0.01％～30％、0.01％～29％、0.01％～28％、0.01％～27％、0.01％～26％、0.01％～25％、0.01％～24％、0.01％～23％、0.01％～22％、0.01％～21％、0.01％～20％、0.01％～19％、0.01％～18％、0.01％～17％、0.01％～16％、0.01％～15％、0.01％～14％、0.01％～13％、0.01％～12％、0.01％～11％、0.01％～10％、0.01％～9.5％、0.01％～9％、0.01％～8.5％、0.01％～8％、0.01％～7.5％、0.01％～7％、0.01％～6.5％、0.01％～6％、0.01％～5.5％、0.01％～5％、0.01％～4.5％、0.01％～4％、0.01％～3.5％、0.01％～3％ or 0.01 to 2.5%.

The elongation coefficient of the multifilament yarn may be 0.1％～32％、0.1％～31％、0.1％～30％、0.1％～29％、0.1％～28％、0.1％～27％、0.1％～26％、0.1％～25％、0.1％～24％、0.1％～23％、0.1％～22％、0.1％～21％、0.1％～20％、0.1％～19％、0.1％～18％、0.1％～17％、0.1％～16％、0.1％～15％、0.1％～14.5％、0.1％～14％、0.1％～13.5％、0.1％～13％、0.1％～12.5％、0.1％～12％、0.1％～11.5％、0.1％～11％、0.1％～10.5％、0.1％～10％、0.1％～9.5％、0.1％～9％、0.1％～8.5％、0.1％～8％、0.1％～7.5％、0.1％～7％、0.1％～6.5％、0.1％～6％、0.1％～5.5％、0.1％～5％、0.1％～4.5％、0.1％～4％、0.1％～3.5％、0.1％～3％ or 0.1% to 2.5%, or 0.5% to less than 33％、0.5％～32％、0.5％～31％、0.5％～30％、0.5％～29％、0.5％～28％、0.5％～27％、0.5％～26％、0.5％～25％、0.5％～24％、0.5％～23％、0.5％～22％、0.5％～21％、0.5％～20％、0.5％～19％、0.5％～18％、0.5％～17％、0.5％～16％、0.5％～15％、0.5％～14.5％、0.5％～14％、0.5％～13.5％、0.5％～13％、0.5％～12.5％、0.5％～12％、0.5％～11.5％、0.5％～10％、0.5％～9.5％、0.5％～9％、0.5％～8.5％、0.5％～8％、0.5％～7.5％、0.5％～7％、0.5％～6.5％、0.5％～6％、0.5％～5.5％、0.5％～5％、0.5％～4.5％、0.5％～4％、0.5％～3.5％、0.5％～3％ or 0.5% to 2.5%, or 0.8% to less than 33％、0.8％～32％、0.8％～31％、0.8％～30％、0.8％～29％、0.8％～28％、0.8％～27％、0.8％～26％、0.8％～25％、0.8％～24％、0.8％～23％、0.8％～22％、0.8％～21％、0.8％～20％、0.8％～19％、0.8％～18％、0.8％～17％、0.8％～16％、0.8％～15％、0.8％～14.5％、0.8％～14％、0.8％～13.5％、0.8％～13％、0.8％～12.5％、0.8％～12％、0.8％～11.5％、0.8％～10％、0.8％～9.5％、0.8％～9％、0.8％～8.5％、0.8％～8％、0.8％～7.5％、0.8％～7％、0.8％～6.5％、0.8％～6％ or 0.8% to 5.5%, or 1% to less than 33％、1％～32％、1％～31％、1％～30％、1％～29％、1％～28％、1％～27％、1％～26％1％～25％、1％～24％、1％～23％、1％～22％、1％～21％、1％～20％、1％～19％、1％～18％、1％～17％、1％～16％、1％～15％、1％～14.5％、1％～14％、1％～13.5％、1％～13％、1％～12％、1％～12％、1％～11.5％、1％～10％、1％～9％、1％～8％、1％～7％、1％～6.5％、1％～6％、1％～5.5％、1％～5％ or 1% to 4%, or 1.5% to less 33％、1.5％～32％、1.5％～31％、1.5％～30％、1.5％～29％、1.5％～28％、1.5％～27％、1.5％～26％、1.5％～25％、1.5％～24％、1.5％～23％、1.5％～22％、1.5％～21％、1.5％～20％、1.5％～19％、1.5％～18％、1.5％～17％、1.5％～16％、1.5％～15％、1.5％～14.5％、1.5％～14％、1.5％～13.5％、1.5％～13％、1.5％～12％、1.5％～12％、1.5％～11.5％、1.5％～10％、1.5％～9％、1.5％～8％、1.5％～7％、1.5％～6.5％、1.5％～6％、1.5％～5.5％ or 1.5% to 5%, or 2% to less 33％、2％～32％、2％～31％、2％～30％、2％～29％、2％～28％、2％～27％、2％～26％、2％～25％、2％～24％、2％～23％、2％～22％、2％～21％、2％～20％、2％～19％、2％～18％、2％～17％、2％～16％、2％～15％、2％～14.5％、2％～14％、2％～13.5％、2％～13％、2％～12％、2％～12％、2％～11.5％、2％～10％、2％～9％、2％～8％、2％～7％、2％～6.5％、2％～6％、2％～5.5％、2％～5％ or 2% to 4%, or 2.5% to 10%, 2.5% to 9%, 2.5% to 8%, 2.5% to 7%, 2.5% to 6%, 2.5% to 5.5%, 2.5% to 5%, 2.5% to 4.5% or 2.5% to 4%.

The elongation of the multifilament may be appropriately selected depending on the application, and may be, for example, 1% to 100%, 3％～100％、5％～100％、5％～95％、5％～90％、5％～85％、5％～80％、5％～70％、5％～65％、5％～60％、5％～55％、5％～50％、8％～100％、8％～95％、8％～90％、8％～85％、8％～80％、8％～75％、8％～70％、8％～65％、8％～60％、8％～55％、8％～50％、10％～100％、10％～95％、10％～90％、10％～85％、10％～80％、10％～75％、10％～70％、10％～65％、10％～60％、10％～55％%, or 10% to 50%.

The upper limit of the coefficient of variation in the strength of the multifilament is more preferably 20% or less. When the coefficient of variation in the strength of the multifilament is 20% or less, the process passability in the post-process steps such as twisting, spinning, weaving, braiding, cutting and the like can be further improved. The lower limit value of the coefficient of variation of the strength may be 0.01% or more, 0.1% or more, 0.2% or more, 0.3% or more, 0.4% or more, 0.5% or more, or 0.6% or more, and may be appropriately selected in consideration of productivity. The coefficient of variation in the strength of the multifilament yarn is preferably 19% or less, 18% or less, 17% or less, 16% or less, 15% or less, 14.5% or less, 14% or less, 13.5% or less, 13% or less, 12.5% or less, 12% or less, 11.5% or less, or 11% or less, more preferably 10.5% or less, 10% or less, 9.5% or less, 9% or less, 8.5% or less, 8% or less, 7.5% or less, 7% or less, 6.5% or less, 6% or less, 5.5% or less, 5% or less, 4.5% or less, 4% or less, 3.5% or less, 3% or 2.5% or less.

Further, it is preferably 0.01% to less than 0.1%, more than 0.1% to less than 1%, more than 1% to less than 5%, more than 5% to less than 10% or more than 10% to less than 15%, more preferably 0.01% to less than 0.1%, more than 1% to less than 5% or more than 5% to less than 10%, more preferably 0.01% to less than 0.1%, more than 0.1% to less than 1% or more than 1% to less than 5%, still more preferably 0.01% to less than 0.1%, more than 0.1% to less than 1% or more than 1% to 3.8%, particularly preferably 0.01% to less than 0.1%, more than 0.1% to less than 1% or more than 1% to less than 3.5%. The coefficient of variation in the strength of the multifilament yarn is preferably 0.01％～20％、0.01％～19％、0.01％～18％、0.01％～17％、0.01％～16％、0.01％～15％、0.01％～14.5％、0.01％～14％、0.01％～13.5％、0.01％～13％、0.01％～12.5％、0.01％～12％、0.01％～11.5％、0.01％～11％、0.01％～10.5％、0.01％～10％、0.01％～9.5％、0.01％～9％、0.01％～8.5％、0.01％～8％、0.01％～7.5％、0.01％～7％、0.01％～6.5％、0.01％～6％、0.01％～5.5％、0.01％～5％、0.01％～4.5％、0.01％～4％、0.01％～3.8％、0.01％～3.5％、0.01％～3％、0.01％～2.5％、0.01％～2％、0.01％～1.5％ or 0.01% to 1%, 0.1％～20％、0.1％～19％、0.1％～18％、0.1％～17％、0.0.1％～16％、0.1％～15％、0.1％～14.5％、0.1％～14％、0.1％～13.5％、0.1％～13％、0.1％～12.5％、0.1％～12％、0.1％～11.5％、0.1％～11％、0.1％～10.5％、0.1％～10％、0.1％～9.5％、0.1％～9％、0.1％～8.5％、0.1％～8％、0.1％～7.5％、0.1％～7％、0.1％～6.5％、0.1％～6％、0.1％～5.5％、0.1％～5％、0.1％～4.5％、0.1％～4％、0.1％～3.5％、0.1％～3％、0.1％～2.5％、0.1％～2％、0.1％～1.5％ or 0.1% to 1%, 0.2％～20％、0.2％～19％、0.2％～18％、0.2％～17％、0.0.2％～16％、0.2％～15％、0.2％～14.5％、0.2％～14％、0.2％～13.5％、0.2％～13％、0.2％～12.5％、0.2％～12％、0.2％～11.5％、0.2％～11％、0.2％～10.5％、0.2％～10％、0.2％～9.5％、0.2％～9％、0.2％～8.5％、0.2％～8％、0.2％～7.5％、0.2％～7％、0.2％～6.5％、0.2％～6％、0.2％～5.5％、0.2％～5％、0.2％～4.5％、0.2％～4％、0.2％～3.5％、0.2％～3％ or 0.2% to 2.5%, 0.3％～20％、0.3％～19％、0.3％～18％、0.3％～17％、0.0.3％～16％、0.3％～15％、0.3％～14.5％、0.3％～14％、0.3％～13.5％、0.3％～13％、0.3％～12.5％、0.3％～12％、0.3％～11.5％、0.3％～11％、0.3％～10.5％、0.3％～10％、0.3％～9.5％、0.3％～9％、0.3％～8.5％、0.3％～8％、0.3％～7.5％、0.3％～7％、0.3％～6.5％、0.3％～6％、0.3％～5.5％、0.3％～5％、0.3％～4.5％、0.3％～4％、0.3％～3.5％、0.3％～3％ or 0.3% to 2.5%, 0.4％～20％、0.4％～19％、0.4％～18％、0.4％～17％、0.0.4％～16％、0.4％～15％、0.4％～14.5％、0.4％～14％、0.4％～13.5％、0.4％～13％、0.4％～12.5％、0.4％～12％、0.4％～11.5％、0.4％～11％、0.4％～10.5％、0.4％～10％、0.4％～9.5％、0.4％～9％、0.4％～8.5％、0.4％～8％、0.4％～7.5％、0.4％～7％、0.4％～6.5％、0.4％～6％、0.4％～5.5％、0.4％～5％、0.4％～4.5％、0.4％～4％、0.4％～3.5％、0.4％～3％ or 0.4% to 2.5%, 0.5％～20％、0.5％～19％、0.5％～18％、0.5％～17％、0.0.5％～16％、0.5％～15％、0.5％～14.5％、0.5％～14％、0.5％～13.5％、0.5％～13％、0.5％～12.5％、0.5％～12％、0.5％～11.5％、0.5％～11％、0.5％～10.5％、0.5％～10％、0.5％～9.5％、0.5％～9％、0.5％～8.5％、0.5％～8％、0.5％～7.5％、0.5％～7％、0.5％～6.5％、0.5％～6％、0.5％～5.5％、0.5％～5％、0.5％～4.5％、0.5％～4％、0.5％～3.5％、0.5％～3％ or 0.5% to 2.5%.

The upper limit of the coefficient of variation of the elastic modulus of the modified fibroin multifilament is preferably 20% or less. When the coefficient of variation of the elastic modulus is 20% or less, the process passability in the post-process steps such as twisting, spinning, weaving, braiding, cutting and the like can be further improved. The lower limit value of the coefficient of variation of the elastic modulus of the multifilament may be 0.01% or more, 0.1% or more, 0.2% or more, 0.3% or more, 0.4% or more, 0.5% or more, 0.6% or more, 0.7% or more, 0.8% or more, 0.9% or more, or 1% or more, and may be appropriately selected in consideration of productivity. The coefficient of variation in the elastic modulus of the multifilament is preferably 19% or less, 18% or less, 17% or less, 16% or less, 15% or less, 14.5% or less, 14% or less, 13.5% or less, 13% or less, 12.5% or less, 12% or less, 11.5% or less, or 11% or less, more preferably 10.5% or less, 10% or less, less than 10%, 9.5% or less, 9% or less, 8.5% or less, 8% or less, 7.5% or less, 7% or less, 6.5% or less, 6% or less, 5.5% or 5% or less.

Further, the coefficient of variation of the elastic modulus of the multifilament is preferably 0.01% to less than 0.1%, more than 0.1% to less than 1%, more than 1% to less than 5%, more than 5% to less than 10% or more than 10% to less than 15%, more preferably 0.01% to less than 0.1%, more preferably 0.1% to less than 1%, more preferably 1% to less than 5% or more than 5% to less than 10%, more preferably 0.01% to less than 0.1%, more preferably 0.1% to less than 1% or more than 1% to less than 5%. The modulus of elasticity of the multifilament yarn is preferably 0.01％～20％、0.01％～19％、0.01％～18％、0.01％～17％、0.01％～16％、0.01％～15％、0.01％～14.5％、0.01％～14％、0.01％～13.5％、0.01％～13％、0.01％～12.5％、0.01％～12％、0.01％～11.5％、0.01％～11％、0.01％～10.5％、0.01％～10％、0.01％～9.5％、0.01％～9％、0.01％～8.5％、0.01％～8％、0.01％～7.5％、0.01％～7％、0.01％～6.5％、0.01％～6％、0.01％～5.5％、0.01％～5％、0.01％～4.5％、0.01％～4％、0.01％～3.5％、0.01％～3％、0.01％～2.5％ or 0.01 to 2%, 0.1％～20％、0.1％～19％、0.1％～18％、0.1％～17％、0.0.1％～16％、0.1％～15％、0.1％～14.5％、0.1％～14％、0.1％～13.5％、0.1％～13％、0.1％～12.5％、0.1％～12％、0.1％～11.5％、0.1％～11％、0.1％～10.5％、0.1％～10％、0.1％～9.5％、0.1％～9％、0.1％～8.5％、0.1％～8％、0.1％～7.5％、0.1％～7％、0.1％～6.5％、0.1％～6％、0.1％～5.5％、0.1％～5％、0.1％～4.5％、0.1％～4％、0.1％～3.5％、0.1％～3％、0.1％～2.5％、0.1％～2％,, 0.2％～20％、0.2％～19％、0.2％～18％、0.2％～17％、0.2％～16％、0.2％～15％、0.2％～14.5％、0.2％～14％、0.2％～13.5％、0.2％～13％、0.2％～12.5％、0.2％～12％、0.2％～11.5％、0.2％～11％、0.2％～10.5、0.2％～10％、0.2％～9.5％、0.2％～9％、0.2％～8.5％、0.2％～8％、0.2％～7.5％、0.2％～7％、0.2％～6.5％、0.2％～6％、0.2％～5.5％、0.2％～5％、0.2％～4.5％、0.2％～4％、0.2％～3.5％、0.2％～3％ or 0.2 to 2.5%, 0.3％～20％、0.3％～19％、0.3％～18％、0.3％～17％、0.3％～16％、0.3％～15％、0.3％～14.5％、0.3％～14％、0.3％～13.5％、0.3％～13％、0.3％～12.5％、0.3％～12％、0.3％～11.5％、0.3％～11％、0.3％～10.5％、0.3％～10％、0.3％～9.5％、0.3％～9％、0.3％～8.5％、0.3％～8％、0.3％～7.5％、0.3％～7％、0.3％～6.5％、0.3％～6％、0.3％～5.5％、0.3％～5％、0.3％～4.5％、0.3％～4％、0.3％～3.5％、0.3％～3％ or 0.3 to 2.5%, 0.4％～20％、0.4％～19％、0.4％～18％、0.4％～17％、0.4％～16％、0.4％～15％、0.4％～14.5％、0.4％～14％、0.4％～13.5％、0.4％～13％、0.4％～12.5％、0.4％～12％、0.4％～11.5％、0.4％～11％、0.4％～10.5％、0.4％～10％、0.4％～9.5％、0.4％～9％、0.4％～8.5％、0.4％～8％、0.4％～7.5％、0.4％～7％、0.4％～6.5％、0.4％～6％、0.4％～5.5％、0.4％～5％、0.4％～4.5％、0.4％～4％、0.4％～3.5％、0.4％～3％ or 0.4 to 2.5%, 0.5％～20％、0.5％～19％、0.5％～18％、0.5％～17％、0.0.5％～16％、0.5％～15％、0.5％～14.5％、0.5％～14％、0.5％～13.5％、0.5％～13％、0.5％～12.5％、0.5％～12％、0.5％～11.5％、0.5％～11％、0.5％～10.5％、0.5％～10％、0.5％～9.5％、0.5％～9％、0.5％～8.5％、0.5％～8％、0.5％～7.5％、0.5％～7％、0.5％～6.5％、0.5％～6％、0.5％～5.5％、0.5％～5％、0.5％～4.5％ or 0.5 to 4%, 1 to 10%, 1 to 9%, 1 to 8%, 1 to 7%, 1 to 6%, 1 to 5% or 1 to 4%.

The upper limit of the coefficient of variation of the fineness of the multifilament is preferably 20% or less. When the coefficient of variation of the fineness is 20% or less, the process passability in the post-process steps such as twisting, spinning, weaving, knitting, cutting and the like can be further improved. The lower limit of the coefficient of variation of the fineness of the multifilament may be 0.01% or more, 0.1% or more, 0.2% or more, 0.3% or more, or 0.4% or more, and may be appropriately selected in consideration of productivity. The coefficient of variation of the fineness of the multifilament is preferably 19% or less, 18% or less, 17% or less, 16% or less, 15% or less, 14% or less, 13% or less, 12% or less, or 11% or less, more preferably 10% or less, 9% or less, 8% or less, 7% or less, 6% or 5% or less. Further, the coefficient of variation of the fineness of the multifilament yarn is preferably 0.01% to less than 0.1%, more than 0.1% to less than 1%, more than 1% to less than 5%, more than 5% to less than 10%, more than 10% to less than 15% or more than 15% to less than 20%, more preferably 0.01% to less than 0.1%, more than 0.1% to less than 1%, more preferably 0.01% to less than 5%, more preferably 0.1% to less than 1%, more than 1% to less than 5% or more than 5% to less than 10%, more preferably 0.01% to less than 0.1%, more than 0.1% to less than 1%, more preferably more than 1% to less than 5% or more than 5% to 6.7%, even more preferably 0.01% to less than 0.1%, more than 1% to less than 1% or more than 1% and particularly preferably 0.01% to less than 10% to less than 15%, more preferably 0.01% to less than 1%, more than 1% to less than 1% and more preferably 0.3% to less than 3%.

The coefficient of variation of the fineness of the multifilament is preferably 0.01％～15％、0.01％～14％、0.01％～13％、0.01％～12％、0.01％～11％、0.01％～10％、0.01％～9.5％、0.01％～9％、0.01％～8.5％、0.01％～8％、0.01％～7.5％、0.01％～7％、0.01％～6.7％、0.01％～6.5％、0.01％～6％、0.01％～5.5％、0.01％～5％、0.01％～4.5％、0.01％～4％、0.01％～3.5％、0.01％～3％、0.01％～2.5％、0.01％～2％、0.01％～1.5％、0.01％～1％., and the coefficient of variation of the fineness of the multifilament may be 0.1％～20％、0.1％～19％、0.1％～18％、0.1％～17％、0.1％～16％、0.1％～15％、0.1％～14％、0.1％～13％、0.1％～12％、0.1％～11％、0.1％～10％、0.1％～9.5％、0.1％～9％、0.1％～8.5％、0.1％～8％、0.1％～7.5％、0.1％～7％、0.1％～6.5％、0.1％～6％、0.1％～5.5％、0.1％～5％、0.1％～4.5％、0.1％～4％、0.1％～3.5％、0.1％～3％、0.1％～2.5％、0.1％～2％、0.1％～1.5％ or 0.1% to 1%, or may be 0.2％～20％、0.2％～19％、0.2％～18％、0.2％～17％、0.2％～16％、0.2％～15％、0.2％～14％、0.2％～13％、0.2％～12％、0.2％～11％、0.2％～10％、0.2％～9.5％、0.2％～9％、0.2％～8.5％、0.2％～8％、0.2％～7.5％、0.2％～7％、0.2％～6.5％、0.2％～6％、0.2％～5.5％、0.2％～5％、0.2％～4.5％、0.2％～4％、0.2％～3.5％、0.2％～3％ or 0.2% to 2.5%, or may be 0.3％～20％、0.3％～19％、0.3％～18％、0.3％～17％、0.3％～16％、0.3％～15％、0.3％～14％、0.3％～13％、0.3％～12％、0.3％～11％、0.3％～10％、0.3％～9.5％、0.3％～9％、0.3％～8.5％、0.3％～8％、0.3％～7.5％、0.3％～7％、0.3％～6.5％、0.3％～6％、0.3％～5.5％、0.3％～5％、0.3％～4.5％、0.3％～4％、0.3％～3.5％、0.3％～3％ or 0.3% to 2.5%, or may be 0.4％～20％、0.4％～19％、0.4％～18％、0.4％～17％、0.4％～16％、0.4％～15％、0.5％～14％、0.4％～13％、0.4％～12％、0.4％～11％、0.4％～10％、0.4％～9.5％、0.4％～9％、0.4％～8.5％、0.4％～8％、0.4％～7.5％、0.4％～7％、0.4％～6.5％、0.4％～6％、0.4％～5.5％、0.4％～5％、0.4％～4.5％、0.4％～4％、0.4％～3.5％ or 0.4% to 3%, or 0.4% to 2.5%, or may be 0.5％～20％、0.5％～19％、0.5％～18％、0.5％～17％、0.5％～16％、0.5％～15％、0.5％～14％、0.5％～13％、0.5％～12％、0.5％～11％、0.5％～10％、0.5％～9.5％、0.5％～9％、0.5％～8.5％、0.5％～8％、0.5％～7.5％、0.5％～7％、0.5％～6.5％、0.5％～6％、0.5％～5.5％、0.5％～5％、0.5％～4.5％、0.5％～4％、0.5％～3.5％、0.5％～3％ or 0.5% to 2.5%, or may be 0.3% to 3%, or 0.3% to 2.8%.

The value of the fineness [ D ] ([ den ]) of the multifilament may be appropriately selected depending on the application, and for example, the fineness ([ D/filement ]) of each monofilament may be 0.7 to 150D, 0.7～140、0.7～130D、0.7～120D、0.7～110D、0.8～100D、0.7～100D、0.7～90D、0.7～80D、0.7～70D、0.7～60D、0.7～50D、0.7～40D、0.7～30D、0.7～20D、0.7～15D、0.7～10D、0.7～9D、0.7～8D、0.7～7D、0.7～6D、0.7～5D、0.7～4D、0.7～3D、0.7～2.5D、0.7～2.2D、0.7～2D、0.7～1.8D、0.7～1.6D、0.7～1.5D、0.7～1.4D、0.7～1.3D、0.7～1.2D、0.7～1.1D or 0.7 to 1D, or may be 0.8 to 3D, 0.8 to 2.5D, 0.8 to 2.2D, 0.8 to 2D, 0.8 to 1.8D, 0.8 to 1.6D, 0.8 to 1.5D, 0.8 to 1.4D, 0.8 to 1.3D, 0.8 to 1.2D, 0.8 to 1.1D or 0.8 to 1D, or may be 1 to 3D, 1 to 2.5D, 1 to 2.2D, 1 to 2D, 1 to 1.9D, 1 to 1.8D, 1 to 1.7D or 1 to 1.6D. The density of the recombinant structural protein may be, for example, 1.3 to 1.4[ g/cm ³ ], or 1.35[ g/cm ³ ] or 1.34[ g/cm ³ ].

The multifilament according to the present embodiment may have a shrinkage history in which irreversible shrinkage occurs after spinning. The shrinkage history is a shrinkage history in which irreversible shrinkage occurs by contacting the multifilament yarn with water, and/or a shrinkage history in which irreversible shrinkage occurs by heat-relaxing the multifilament yarn. The multifilament according to the present embodiment is, for example, a multifilament obtained by the above-described production method, and therefore is a multifilament containing substantially no residual stress due to stretching during spinning.

< Shrinkage ratio >

The multifilament yarn according to the present embodiment having a shrinkage history in which irreversible shrinkage occurs after spinning preferably has a shrinkage ratio defined by the following formula of 5% or less. The multifilament in the following formula is a multifilament having a shrinkage history in which irreversible shrinkage occurs after spinning.

The shrinkage of the fiber caused by contact with moisture can be evaluated, for example, by using the shrinkage ratio obtained by the above formula as an index. The "length of the multifilament yarn in a wet state" and the "length of the multifilament yarn in a dry state from a wet state" can be measured, for example, by the following method.

The multifilament yarn was cut into a length of about 30cm to prepare a fiber bundle. The fiber bundles were immersed (wetted) in water at 40 ℃ for 15 minutes and dried at room temperature for 2 hours. After drying, the length of the fiber bundle was measured. The wet and dry are repeated at least 3 times again, and the average length in wet state is defined as "length of multifilament in wet state" and the average length in dry state is defined as "length of multifilament in dry state from wet state".

In the multifilament, the smaller the shrinkage, the better, and in particular, in the article such as a fabric made of multifilament, the smaller the shrinkage portion is preferable.

The shrinkage defined by the above formula is preferably 5.0% or less, but may be 4.5% or less, 4% or less, 3.5% or less, 3.2% or less, 3.1% or less, 3.0% or less, 2.9% or less, 2.8% or less, 2.7% or less, 2.6% or less, 2.5% or less, 2.4% or less, 2.3% or less, 2.2% or less, 2.1% or less, 2.0% or less, 1.9% or less, 1.8% or less, 1.7% or less, 1.6% or less, 1.5% or less, 1.4% or less, 1.3% or less, 1.2% or less, 1.1% or less, 1.0% or less, 0.9% or 0.8% or less, 0.7% or less, 0.6% or less, 0.5% or less.

(Product)

The modified fibroin multifilament according to the present embodiment can be applied as a fiber or yarn (spun yarn, twisted yarn, false twisted yarn, processed yarn, mixed yarn, blended yarn, or the like) to woven fabrics (fabrics), knitted, woven, nonwoven fabrics, paper, cotton, or the like. Further, the present invention can be applied to high-strength applications such as wire loops, surgical sutures, flexible handles for electrical components, and physiologically active materials for implantation (e.g., artificial ligaments and aortic ribbons). These can be manufactured according to a known method.

The modified fibroin fibers may have a critical oxygen index (LOI) value of 18 or more, 20 or more, 22 or more, 24 or more, 26 or more, 28 or more, 29 or more, or 30 or more. The LOI value is measured by a test method of powdery or low melting point synthetic resin according to Japanese national fire protection agency hazard regulation No. section chief, no. 5, no. 31 and 1995.

The modified fibroin fiber may have a maximum moisture absorption and heat generation degree of more than 0.025 ℃/g, 0.026 ℃/g or more, 0.027 ℃/g or more, 0.028 ℃/g or more, 0.029 ℃/g or more, 0.030 ℃/g or more, 0.035 ℃/g or more, or 0.040 ℃/g or more, as determined by the following formula A. The upper limit of the maximum hygroscopic heat generation degree is not particularly limited, and is usually 0.060 ℃/g or less.

Formula A: maximum hygroscopic heat generation = { (maximum value of sample temperature when the sample is moved to high humidity after the sample is placed in low humidity until the sample temperature reaches equilibrium) - (sample temperature when the sample is placed in low humidity until the sample temperature reaches equilibrium) } (°c)/sample weight (g) when the sample is moved to high humidity

The modified fibroin fiber is preferably a fiber having excellent heat insulating properties, and the heat insulating property index obtained from the following formula C may be 0.20 or more.

Formula C: thermal insulation performance index = thermal insulation (%)/weight per unit area of sample (g/m ²)

The modified fibroin fiber may have a thermal insulation performance index of 0.22 or more, 0.24 or more, 0.26 or more, 0.28 or more, 0.30 or more, or 0.32 or more. The upper limit of the heat insulating property index is not particularly limited, and may be, for example, 0.60 or less or 0.40 or less.

Examples

[ Production of modified fibroin ] (1) preparation of expression vector

Modified spider silk fibroin having a sequence number 40 (hereinafter also referred to as "PRT 966") and modified spider silk fibroin having a sequence number 15 (hereinafter also referred to as "PRT 799") were designed based on the base sequence and amino acid sequence of fibroin (GenBank accession numbers: P46804.1, GI: 1174415) from Jin Fang spiders (NEPHILA CLAVIPES). In addition, the amino acid sequence shown in SEQ ID NO. 40 has a sequence in which all of QQQs in a sequence in which 20 domain sequence regions existing in the amino acid sequence shown in SEQ ID NO. 7 are repeated 2 times are replaced with VF and the remaining Qs are replaced with I, and further the amino acid sequence shown in SEQ ID NO. 11 (tag sequence and hinge sequence) is added to the N-terminus. The amino acid sequence shown in SEQ ID NO. 15 has an amino acid sequence obtained by performing substitution, insertion and deletion of an amino acid residue with respect to a silk fibroin amino acid sequence derived from Jin Fang spiders for the purpose of improving productivity, and further the amino acid sequence shown in SEQ ID NO. 11 (tag sequence and hinge sequence) is added to the N-terminus.

Next, nucleic acids encoding artificial structural proteins (engineered fibroin) PRT966 and PRT799 having the amino acid sequences of SEQ ID Nos. 40 and 15 were synthesized. In this nucleic acid, an NdeI site was added to the 5' -end, and an EcoRI site was added downstream of the stop codon. The nucleic acid was cloned into a cloning vector (pUC 118). Then, the nucleic acid was subjected to restriction enzyme treatment with NdeI and EcoRI, and then, the resultant was recombined into a protein expression vector pET-22b (+), respectively, to obtain an expression vector.

(2) Engineering expression of fibroin

Coli BLR (DE 3) was transformed with the expression vector obtained in (1). The transformed E.coli was cultured in 2mL of LB medium containing ampicillin for 15 hours. This culture broth was added to 100mL of seed culture medium (Table 4) containing ampicillin so that the OD ₆₀₀ was 0.005. The culture broth temperature was kept at 30℃and flask culture was performed until OD ₆₀₀ reached 5 (about 15 hours), to obtain a seed culture broth.

TABLE 4

Culture medium for seed culture

The seed culture was added to a fermenter with 500mL of production medium (Table 5) to bring the OD ₆₀₀ to 0.05. The culture was carried out while maintaining the temperature of the culture at 37℃and constantly controlling the pH at 6.9. In addition, the dissolved oxygen concentration in the culture broth was maintained at 20% of the dissolved oxygen saturation concentration.

TABLE 5

Production medium

Immediately after complete consumption of glucose in the production medium, the feed solution (455 g/1L glucose, 120g/1L yeast extract) was added at a rate of 1 mL/min. The culture was carried out while maintaining the temperature of the culture at 37℃and constantly controlling the pH at 6.9. In addition, the concentration of dissolved oxygen in the culture solution was maintained at 20% of the saturation concentration of dissolved oxygen, and the culture was continued for 20 hours. Then, 1M isopropyl- β -thiogalactopyranoside (IPTG) was added to the culture broth to a final concentration of 1mM, and the induced remodelling fibroin was expressed. After the lapse of 20 hours from the addition of IPTG, the culture medium was centrifuged to collect the cells. SDS-PAGE was performed using cells prepared from the culture medium before and after addition of IPTG, and expression of the target modified fibroin was confirmed based on the occurrence of bands depending on the size of the target modified fibroin added by IPTG.

(3) Modified fibroin purification

The cells recovered 2 hours after the addition of IPTG were washed with 20mM Tris-HCl buffer (pH 7.4). The washed cells were suspended in 20mM Tris-HCl buffer (pH 7.4) containing about 1mM PMSF, and the cells were disrupted by a high-pressure homogenizer (manufactured by GEA Niro Soavi Co.). And (3) centrifuging the crushed cells to obtain a precipitate. The resulting precipitate was washed with 20mM Tris-HCl buffer (pH 7.4) until high purity was achieved. The washed precipitate was suspended in 8M guanidine buffer (8M guanidine hydrochloride, 10mM sodium dihydrogen phosphate, 20mM NaCl, 1mM Tris-HCl, pH 7.0) so as to reach a concentration of 100mg/mL, and stirred at 60℃for 30 minutes with a stirrer to dissolve the precipitate. After dissolution, the solution was dialyzed with water using a dialysis tube (cellulose tube 36/32 manufactured by Sanguang pure chemical Co., ltd.). The white coagulated protein obtained after dialysis was recovered by centrifugation, the water was removed by a freeze-dryer, and the freeze-dried powder was recovered to obtain modified fibroin (PRT 966 and PRT 799).

(Production of modified fibroin multifilament)

(1) Preparation of spinning dope (dope)

26 Mass% of modified fibroin (PRT 966) obtained in the modified fibroin production process described above and 74 mass% of formic acid (purity 98% manufactured by the chemical company, korea, co.) as a solvent for dissolution were mixed and heated with stirring for 1 hour with an aluminum block heater at 40 ℃ to dissolve the modified fibroin. Filtering and defoaming by using a metal filter with meshes of 1 mu m to obtain spinning solution.

(2) Dry and wet spinning

Example 1

Spinning was performed using a spinning device shown in fig. 6. The prepared dope was filled into a liquid tank (a reservoir), and the dope was ejected from a spinning nozzle (a spinning nozzle) having 100 holes through an air gap into a coagulation bath using a gear pump to form a dope. Next, the coagulated filaments were drawn in an aqueous wash bath. After washing and stretching in a water washing bath, the obtained modified fibroin multifilament was dried using a dry heat plate and wound up with a winder. The number of the multifilaments was 100. The conditions for dry and wet spinning were as follows.

Pore size of the spinning spinneret: 0.08mm

Number of holes of spinning nozzle: 100

And (3) coagulating liquid: 100% methanol

Temperature of the coagulation liquid: 10 DEG C

Temperature of the water wash bath: 40 DEG C

Temperature of the stretching bath: 40 DEG C

Total stretch ratio: 4.4 times

Drying temperature: 70 DEG C

Example 2

Modified fibroin multifilament yarn was produced in the same manner as in example 1 except that a spinning nozzle having a pore diameter of 0.1mm and a pore number of 360 pores and a total draw ratio of 5.3 times was used. The number of the multifilaments was 360.

(3) Wet spinning

Example 3

Spinning was performed using a spinning device shown in fig. 7. The prepared dope was filled into a liquid tank, and the dope was ejected from a spinning nozzle having 200 holes into a coagulation bath using a gear pump to form a yarn (silk). Next, the coagulated filaments were drawn in an aqueous wash bath. After washing and stretching in a water washing bath, the obtained modified fibroin multifilament was dried using a dry heat plate and wound up with a winder. The number of the multifilaments was 200. Further, the obtained multifilament yarn is formed with recesses extending in the axial direction of the filaments on the surface. The conditions for wet spinning are as follows.

Pore diameter of spinning nozzle: 0.05mm

Number of holes of spinning nozzle: 200

And (3) coagulating liquid: mixed solution of 14.4 mass% sodium sulfate and 65.6 mass% water (80 mass% sodium sulfate aqueous solution of 18 mass%), and 20 mass% formic acid

Temperature of the coagulation liquid: 40 DEG C

Temperature of the water wash bath: 40 DEG C

Temperature of the stretching bath: 60 DEG C

Total stretch ratio: 4.3 times

Drying temperature: 60 DEG C

Example 4

Modified fibroin multifilament yarn was produced in the same manner as in example 3, except that a spinning nozzle having a pore diameter of 0.1mm and a pore number of 1,000 was used and the total draw ratio was 6.2 times. The number of the obtained multifilaments was 1,000. Further, the obtained multifilament yarn was formed with recesses extending in the axial direction of the filaments on the surface (fig. 13).

Example 9

A multifilament was produced by wet spinning in the same manner as in example 3, except that the concentration of the dope (dope) was 31 mass%, and a spinning nozzle having a pore diameter of 0.08mm and a pore number of 1,664 was used and the total draw ratio was 5.7 times. The number of the obtained multifilaments was 1,664.

Example 10

A multifilament was produced by wet spinning in the same manner as in example 3, except that the concentration of the dope (dope) was 31 mass%, and a spinning nozzle having a pore diameter of 0.1mm and a pore number of 2,328 was used and the total draw ratio was 5.7 times. The number of the obtained multifilaments was 2,328.

Example 11

A multifilament was produced by wet spinning in the same manner as in example 3, except that the concentration of the dope (dope) was 31 mass%, and a spinning nozzle having a pore diameter of 0.08mm and a pore number of 3,000 was used and the total draw ratio was 5.7 times. The number of the obtained multifilaments was 3,000. A cross-sectional view of the multifilament yarn is shown in FIG. 14.

(Evaluation of modified fibroin multifilament)

The modified fibroin multifilaments obtained in examples 1 to 4 and examples 9 to 11 were measured for elastic modulus [ gf/D ], strength [ g/D ], elongation at break [% ] and fineness [ D ]. The coefficient of variation of the elastic modulus, the coefficient of variation of the strength, the coefficient of variation of the elongation at break, and the coefficient of variation of the fineness were calculated using the following formulas, respectively.

In addition, the density of the modified fibroin used [ g/cm ³ ] was 1.34[ g/cm ³ ].

The elastic modulus [ gf/D ], the strength [ g/D ], and the elongation at break [% ] of the modified fibroin multifilament were measured and the standard deviation was calculated according to JIS L1013 using a 3345 series tensile tester manufactured by Instron corporation. Examples 1 and 3 were measured using a load cell capacity 10N and examples 2,4 and examples 9 to 11 were measured using a load cell capacity 50N under conditions of a test length of 300mm and a test speed of 300 mm/min at a temperature of 20℃and a relative humidity of 65%. The mass of the multifilament cut into 3m length was measured at 20℃under a relative humidity of 65%, and the fineness [ D ] of the modified fibroin multifilament was calculated by converting the mass into a mass of 9,000 m. The average value of each physical property of the modified fibroin multifilament in examples 1 to 4 was calculated as an average value of the sampling number n=5. The average value of each physical property of the multifilament in example 9 was calculated as an average value of the number of samples n=10. The average value of each physical property of the multifilament in example 10 was calculated as an average value of the number of samples n=15. The average value of each physical property of the multifilament in example 11 was calculated as an average value of the number of samples n=30. The values of the respective coefficients of variation were calculated from the above formulas using the respective average values of the measured physical properties of the multifilament (average value of the elastic modulus, average value of the strength, average value of the elongation and average value of the fineness). The evaluation results of the respective coefficients of variation are shown in table 6. The average values and standard deviations of the elongation at break [% ] and the fineness [ D ] of examples 1 to 4 and 9 to 11 are shown in Table 7.

TABLE 6

As shown in Table 6, the productivity was remarkably improved by using the spinning nozzle having the number of holes of 100 to 3,000. Further, the obtained multifilaments (examples 1 to 4 and examples 9 to 11) had a Coefficient of Variation (CV) of elastic modulus of 1.2 to 4.7, a Coefficient of Variation (CV) of strength of 0.8 to 2.2, a Coefficient of Variation (CV) of elongation of 2.6 to 5.3, a Coefficient of Variation (CV) of fineness of 0.04 to 2.6, and extremely small variation in the physical properties of the multifilaments, and remarkably excellent quality stability.

TABLE 7

Examples 5 to 8

(4) Shrinkage step and drying step

The multifilament yarn comprising the modified fibroin obtained in examples 1 to 4 was cut to a length of about 30cm, and each multifilament bundle was immersed in water at 40 ℃ for 90 seconds to shrink. Then, each multifilament bundle was taken out of the water, dried, and the length of each dried multifilament bundle was measured.

(5) Evaluation of shrinkage Property of multifilament yarn with irreversible shrinkage experience

The shrinkage performance of the modified fibroin multifilament obtained in the above (4) can be evaluated using the shrinkage ratio obtained by the following method as an index. Let the number of samples be n=3, and the calculation is performed according to the following equation. The calculation results are shown in Table 8.

TABLE 8

As shown in table 8, the modified fibroin multifilament after the shrinkage treatment had a shrinkage of ±0.4% (examples 5 to 8) or less, and had excellent dimensional stability against moisture. As described above, the multifilament having an irreversible shrinkage history is excellent in dimensional stability against moisture and the relative deviation of the physical properties of the multifilament is extremely small, so that the quality stability is remarkably excellent.

Test examples 1 to 3: stripping Performance test of dope

(1) Preparation of spinning solution

31 Mass% of modified fibroin (PRT 966) obtained in the modified fibroin production process described above and 69 mass% of formic acid (purity 99% by the chemical industry, corporation) as a solvent for dissolution were mixed, and heated with stirring for 1 hour with an aluminum block heater at 40 ℃ to dissolve the modified fibroin. Filtering and defoaming the solution by using a metal filter with a mesh of 1 mu m to prepare spinning solution.

(2) Peeling property test

The stripping performance of the material of the spinning nozzle (spinning nozzle) was evaluated with respect to the spinning liquid. At aboutRespectively dripping spinning solution on the surfaces of round test pieces with different materials. After immersing them in the coagulation liquid for 5 seconds, the dope (semi-coagulated material) was peeled from the surface of each test piece in the coagulation liquid. The stripping performance of the dope was evaluated based on the degree of force required for stripping the dope (semi-coagulated matter). The materials and evaluation results of the spinning nozzles used are shown in Table 9. The coagulation liquid is a mixed solution obtained by mixing an aqueous sodium sulfate solution having a concentration of 18 mass% with formic acid at a ratio of 80 mass% to 20 mass%, respectively. The peeling property was evaluated as follows.

O: good stripping performance (the spinning solution (semi-solidified substance) can be stripped with a small force)

Delta: poor stripping performance (greater force is required to strip the dope (semi-coagulated) than the material evaluated)

TABLE 9

	Spinning nozzle material	Evaluation of dope peelability
			Test example 1	Pt/Au	△
Test example 2	Hastelloy	○
			Test example 3	SUS316L	○

As shown in table 9, when the material of the spinning nozzle was Hastelloy or SUS316L, the dope (semi-coagulated material) could be peeled off from the surface of the test piece with a smaller force than the case where the material of the spinning nozzle was Pt/Au, and it was confirmed that the peeling performance of the dope was better. Thus, the recombinant structural protein has the property of easily binding (adsorbing) Pt/Au. In particular, in the case of using a spinning solution containing modified fibroin, by setting the material of the spinning nozzle to a material (Hastelloy, SUS316L, or the like) from which modified fibroin is easily peeled, clogging of the spinning nozzle due to adhesion of the spinning solution to the spinning nozzle can be further prevented, and the spinning stability can be further improved. It is particularly suitable for mass spinning using a spinning nozzle having a hole number exceeding 100.

Reference example 1: modified fibroin flammability test

The freeze-dried powder of modified fibroin (PRT 799) was added to a dimethyl sulfoxide solution of lithium chloride (concentration: 4.0 mass%) to make the concentration 24 mass%, and mixed using a shaker for 3 hours to dissolve it. Insoluble matter and bubbles were then removed to obtain a modified fibroin solution (spinning dope).

The resulting dope was heated to 90℃and filtered through a metal filter having a mesh size of 5. Mu.m, and then placed in a30 mL stainless steel syringe for standing and deaeration, and then discharged from a solid nozzle having a needle diameter of 0.2mm into a 100 mass% methanol coagulation bath. The ejection temperature was 90 ℃. After coagulation, the obtained filaments are wound up and naturally dried to obtain a modified fibroin fiber (raw material fiber).

Knitted fabrics (thickness: 180 denier, gauge: 18) were produced by circular knitting using a circular knitting machine using twisted filaments obtained by twisting raw material fibers together. The resultant knitted fabric was cut out to 20g and used as a test piece.

The combustibility test is based on the "test method for granular or low melting point synthetic resins" described in "fire department hazard regulation No. 50 (5/31/1995)". The test was carried out at a temperature of 22℃and a relative humidity of 45% and a gas pressure of 1021 hPa. The measurement results (oxygen concentration (%), combustion rate (%), combustion conversion (%)) are shown in table 10.

TABLE 10

Oxygen concentration (%)	Combustion rate (%)	Combustion conversion (%)
			20.0	39.1	40.1
27.0	48.1	49.3
			28.0	51.9	53.2
30.0	53.6	54.9
			50.0	61.2	62.7
70.0	91.1	93.3
			100.0	97.6	100.0

As a result of the combustibility test, the critical oxygen index (LOI) value of the knitted fabric knitted with the modified fibroin (PRT 799) fiber was 27.2. In general, when the LOI value is 26 or more, it is known to have flame retardancy. From this, it was found that the modified fibroin was excellent in flame retardancy.

Reference example 2: moisture absorption and heat generation performance evaluation of modified fibroin

The freeze-dried powder of the modified fibroin was added to a dimethyl sulfoxide solution (concentration: 4.0 mass%) of lithium chloride so that the concentration was 24 mass%, and mixed with a shaker for 3 hours to dissolve it. Insoluble matter and bubbles were then removed to obtain a modified fibroin solution (spinning dope).

The resulting dope was heated to 60℃and filtered through a metal filter having a mesh size of 5. Mu.m, and then placed in a30 mL stainless steel syringe for standing and deaeration, and then discharged from a solid nozzle having a needle diameter of 0.2mm into a 100 mass% methanol coagulation bath. The ejection temperature was 60 ℃. After coagulation, the obtained filaments are wound up and naturally dried to obtain a modified fibroin fiber (raw material fiber).

For comparison, commercially available wool fibers, cotton fibers, tencel fibers, rayon fibers, and polyester fibers were prepared as raw material fibers.

Knitted fabrics are produced by performing a flat knitting process using various raw material fibers by using a flat knitting machine. The thickness and gauge of the knitted fabric using the PRT918 fiber or the PRT799 fiber are shown in table 11. The thickness and gauge of the knitted fabric using the other raw material fibers were adjusted so as to have a coverage factor substantially the same as that of the modified fibroin fiber knitted fabric. Specifically, the following is described.

TABLE 11

Raw material fiber	Thickness [ N ]	Needle pitch [ GG ]
			PRT918	1/30 (Wool yarn count monofilament)	18
PRT799	1/30 (Wool yarn count monofilament)	16
			Wool	2/30 (Double silk)	14
Cotton cotton	2/34 (Double silk)	14
			Tencel	2/30 (Double silk)	15
Artificial fiber	1/38 (Monofilament)	14
			Terylene	1/60 (Monofilament)	14

2 Knitted fabrics cut to 10cm×10cm were bonded together, and four sides were sewn together to prepare test pieces (test specimens). After the test piece was left in a low humidity environment (temperature 20.+ -. 2 ℃ C., relative humidity 40.+ -. 5%) for 4 hours or more, the test piece was moved to a high humidity environment (temperature 20.+ -. 2 ℃ C., relative humidity 90.+ -. 5%) and temperature measurement was performed at 1 minute intervals for 30 minutes by using a temperature sensor mounted at the center inside the test piece.

From the measurement results, the maximum moisture absorption and heat generation were determined according to the following formula a.

Fig. 15 is a graph showing an example of the results of the hygroscopic heat generation performance test. The horizontal axis of the graph indicates the time of placement (minutes) in a high humidity environment, where 0 is the time for the sample to transfer from a low humidity environment to a high humidity environment. The vertical axis of the graph represents the temperature (sample temperature) measured by the temperature sensor. In the graph shown in fig. 15, the point indicated by M corresponds to the highest value of the sample temperature.

The calculation results of the maximum moisture absorption and heat generation degrees of the respective knitted fabrics are shown in table 12.

TABLE 12

Raw material fiber	Maximum moisture absorption and heat generation degree (DEG C/g)
		PRT918	0.040
PRT799	0.031
		Wool	0.020
Cotton cotton	0.021
		Tencel	0.018
Artificial fiber	0.025
		Terylene	0.010

As shown in table 12, it is clear that modified fibroin (PRT 918 and PRT 799) has higher maximum hygroscopic heat generation and excellent hygroscopic heat generation performance than the conventional materials.

Reference example 3: insulation performance assessment of modified fibroin

For comparison, commercially available wool fibers, silk fibers, cotton fibers, rayon fibers, and polyester fibers were prepared as raw material fibers.

Knitted fabrics are produced by performing a flat knitting process using various raw material fibers by using a flat knitting machine. The count, the number of turns, the stitch length, and the weight per unit area of the knitted fabric using the PRT966 fiber or the PRT799 fiber are shown in table 13. The knitted fabric using the other raw material fibers is adjusted so as to have a cover factor substantially the same as that of the knitted fabric of modified fibroin fibers. Specifically, the following is described.

TABLE 13

The thermal insulation performance was evaluated using a KES-F7 Thermolab II tester manufactured by add-on technologies limited and using a dry contact method (a method assuming that skin and clothes are in direct contact in a dry state). A piece of knitted fabric cut into a rectangle of 20 cm. Times.20 cm was used as a test piece (sample). The test piece was set on a hot plate set at a constant temperature (30 ℃ C.) and the amount of heat (a) emitted by the test piece was determined under the condition that the wind speed in the wind tunnel was 30 cm/sec. The amount of heat (B) emitted under the same conditions as those described above was determined without providing the test piece, and the heat retention rate (%) was calculated from the following formula B.

Formula B: heat retention (%) = (1-a/b) ×100

From the measurement results, the thermal insulation performance index was obtained according to the following formula C.

The results of calculation of the heat insulation performance index are shown in Table 14. The higher the thermal insulation performance index of the material is, the more excellent the thermal insulation performance can be estimated.

TABLE 14

Raw material fiber	Thermal insulation performance index
		PRT966	0.33
PRT799	0.22
		Wool	0.16
Silk fabric	0.11
		Cotton cotton	0.13
Artificial fiber	0.02
		Terylene	0.18

As shown in table 14, modified fibroin (PRT 966 and PRT 799) was found to have a higher heat retaining performance index and an excellent heat retaining performance than the conventional materials.

As shown in reference examples 1 to 3, when the modified silk fibroin is modified spider silk fibroin, the heat insulating performance, the hygroscopic and exothermic performance and/or the flame retardancy can be made more excellent. By using the modified spider silk fibroin to form multifilament, multifilament which is excellent in heat insulating property, hygroscopic heat generating property and/or flame retarding property and extremely excellent in quality stability can be obtained.

Symbol description

1. Extrusion device

2. Apparatus for producing undrawn yarn

3. Wet-heat stretching device

4. Drying device

6. Spinning solution

10. Spinning device

20. Coagulation bath

21. Stretching bath

25. Spinning device

36. Multifilament yarn

38. Modified fibroin multifilament yarn

40. Manufacturing apparatus

42. Feeding roller

44. Coiling machine

46. Water bath

48. Drying machine

54. Heater

56. Tensioning roller

58. Hot roller

60. Processing device

62. Drying device

64. Dry-hot plate

140. Relaxation shrinkage unit (heating unit)

141. Delivery unit

142. Coiling unit

146. Speed adjusting unit

147. And a temperature adjusting unit.

Sequence listing

<110> Spiber Co., ltd

<120> Recombinant structural protein multifilament yarn and method for producing the same

<130> 2018PS071WO1

<150> JP2019-021014

<151> 2019-02-07

<150> JP2019-109041

<151> 2019-06-11

<160> 49

<170> Patent In version 3.5

<210> 1

<211> 50

<212> PRT

<213> Spider

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Ser Gly Cys Asp Val Leu Val Gln Ala Leu Leu Glu Val Val Ser Ala

1 5 10 15

Leu Val Ser Ile Leu Gly Ser Ser Ser Ile Gly Gln Ile Asn Tyr Gly

20 25 30

Ala Ser Ala Gln Tyr Thr Gln Met Val Gly Gln Ser Val Ala Gln Ala

35 40 45

Leu Ala

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<210> 2

<211> 30

<212> PRT

<213> Spider

<400> 2

Ser Gly Cys Asp Val Leu Val Gln Ala Leu Leu Glu Val Val Ser Ala

1 5 10 15

Leu Val Ser Ile Leu Gly Ser Ser Ser Ile Gly Gln Ile Asn

20 25 30

<210> 3

<211> 21

<212> PRT

<213> Spider

<400> 3

Ser Gly Cys Asp Val Leu Val Gln Ala Leu Leu Glu Val Val Ser Ala

1 5 10 15

Leu Val Ser Ile Leu

20

<210> 4

<211> 1154

<212> PRT

<213> Artificial sequence

<220>

<223> Recombinant spider silk protein ADF3KAILARGENRSH1

<400> 4

Met His His His His His His His His His His Ser Ser Gly Ser Ser

1 5 10 15

Leu Glu Val Leu Phe Gln Gly Pro Ala Arg Ala Gly Ser Gly Gln Gln

20 25 30

Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly

35 40 45

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly Tyr

50 55 60

Gly Pro Gly Ser Gly Gln Gln Gly Pro Ser Gln Gln Gly Pro Gly Gln

65 70 75 80

Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

85 90 95

Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Ser Gly Gln Gln Gly Pro

100 105 110

Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

115 120 125

Ala Gly Gly Asn Gly Pro Gly Ser Gly Gln Gln Gly Ala Gly Gln Gln

130 135 140

Gly Pro Gly Gln Gln Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala

145 150 155 160

Gly Gly Tyr Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly

165 170 175

Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala

180 185 190

Ala Ala Gly Gly Tyr Gly Pro Gly Ser Gly Gln Gly Pro Gly Gln Gln

195 200 205

Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

210 215 220

Ala Ala Ala Gly Gly Tyr Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

225 230 235 240

Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly

245 250 255

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly

260 265 270

Tyr Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly Gln Gly Pro

275 280 285

Tyr Gly Pro Gly Ala Ser Ala Ala Ser Ala Ala Ser Gly Gly Tyr Gly

290 295 300

Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly Gln

305 310 315 320

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly

325 330 335

Tyr Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly

340 345 350

Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly

355 360 365

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly

370 375 380

Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro

385 390 395 400

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly

405 410 415

Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly

420 425 430

Gln Gly Ala Tyr Gly Pro Gly Ala Ser Ala Ala Ala Gly Ala Ala Gly

435 440 445

Gly Tyr Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro

450 455 460

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly

465 470 475 480

Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly

485 490 495

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly

500 505 510

Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro

515 520 525

Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ala Ser Ala Ala Val Ser

530 535 540

Val Ser Arg Ala Arg Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln

545 550 555 560

Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

565 570 575

Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Ser Gly

580 585 590

Gln Gln Gly Pro Ser Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly

595 600 605

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly

610 615 620

Gly Tyr Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gly Gln Gly Pro

625 630 635 640

Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Gly Gly Asn Gly

645 650 655

Pro Gly Ser Gly Gln Gln Gly Ala Gly Gln Gln Gly Pro Gly Gln Gln

660 665 670

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro

675 680 685

Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly Gln Gly

690 695 700

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly Tyr

705 710 715 720

Gly Pro Gly Ser Gly Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly Gln

725 730 735

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly

740 745 750

Tyr Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly

755 760 765

Gln Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

770 775 780

Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Tyr Gly Gln Gln Gly

785 790 795 800

Pro Gly Gln Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala

805 810 815

Ser Ala Ala Ser Ala Ala Ser Gly Gly Tyr Gly Pro Gly Ser Gly Gln

820 825 830

Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro

835 840 845

Gly Ala Ser Ala Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Ser

850 855 860

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly

865 870 875 880

Gln Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

885 890 895

Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Ser Gly Gln Gln Gly

900 905 910

Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro

915 920 925

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly

930 935 940

Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly Gln Gly Ala Tyr Gly

945 950 955 960

Pro Gly Ala Ser Ala Ala Ala Gly Ala Ala Gly Gly Tyr Gly Pro Gly

965 970 975

Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro

980 985 990

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly

995 1000 1005

Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser

1010 1015 1020

Ala Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Ser Gly Gln

1025 1030 1035

Gln Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Gly

1040 1045 1050

Gln Gly Pro Tyr Gly Pro Gly Ala Ala Ser Ala Ala Val Ser Val

1055 1060 1065

Gly Gly Tyr Gly Pro Gln Ser Ser Ser Val Pro Val Ala Ser Ala

1070 1075 1080

Val Ala Ser Arg Leu Ser Ser Pro Ala Ala Ser Ser Arg Val Ser

1085 1090 1095

Ser Ala Val Ser Ser Leu Val Ser Ser Gly Pro Thr Lys His Ala

1100 1105 1110

Ala Leu Ser Asn Thr Ile Ser Ser Val Val Ser Gln Val Ser Ala

1115 1120 1125

Ser Asn Pro Gly Leu Ser Gly Cys Asp Val Leu Val Gln Ala Leu

1130 1135 1140

Leu Glu Val Val Ser Ala Leu Val Ser Ile Leu

1145 1150

<210> 5

<211> 24

<212> PRT

<213> Artificial sequence

<220>

<223> His tag and start codon

<400> 5

Met His His His His His His His His His His Ser Ser Gly Ser Ser

1 5 10 15

Leu Glu Val Leu Phe Gln Gly Pro

20

<210> 6

<211> 597

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT380

<400> 6

Met Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

20 25 30

Gln Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly

35 40 45

Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro

50 55 60

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala

65 70 75 80

Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Ala Ala

85 90 95

Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln

100 105 110

Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly

115 120 125

Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro

130 135 140

Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

145 150 155 160

Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

165 170 175

Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly

180 185 190

Gln Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly

195 200 205

Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala

210 215 220

Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr

225 230 235 240

Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly

245 250 255

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Tyr Gly Pro

260 265 270

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

275 280 285

Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly

290 295 300

Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro

305 310 315 320

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro

325 330 335

Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Ala Ala Ala

340 345 350

Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala

355 360 365

Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly

370 375 380

Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro

385 390 395 400

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

405 410 415

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

420 425 430

Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala

435 440 445

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Tyr

450 455 460

Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly

465 470 475 480

Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

485 490 495

Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro

500 505 510

Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly

515 520 525

Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser

530 535 540

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala

545 550 555 560

Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly

565 570 575

Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln

580 585 590

Gly Pro Gly Ala Ser

595

<210> 7

<211> 590

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT410

<400> 7

Met Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

20 25 30

Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly

35 40 45

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro

50 55 60

Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

65 70 75 80

Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly Gln

85 90 95

Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

100 105 110

Gly Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala

115 120 125

Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr

130 135 140

Gly Pro Gly Ala Ser Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly

145 150 155 160

Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro

165 170 175

Gly Gln Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

180 185 190

Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly

195 200 205

Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala

210 215 220

Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser

225 230 235 240

Ala Ala Ala Ala Ala Gly Gln Tyr Gly Tyr Gly Pro Gly Gln Gln Gly

245 250 255

Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln

260 265 270

Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

275 280 285

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

290 295 300

Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Tyr Gly

305 310 315 320

Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser

325 330 335

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

340 345 350

Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln

355 360 365

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly

370 375 380

Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

385 390 395 400

Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala

405 410 415

Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr

420 425 430

Gly Pro Gly Gln Ser Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro

435 440 445

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro

450 455 460

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

465 470 475 480

Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly

485 490 495

Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser

500 505 510

Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly

515 520 525

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly

530 535 540

Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser

545 550 555 560

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala

565 570 575

Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser

580 585 590

<210> 8

<211> 565

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT525

<400> 8

Met Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Ala Ala Gly Ser Asn Gly Pro Gly Ser Gly Gln Gln Gly

20 25 30

Pro Gly Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

35 40 45

Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

50 55 60

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala

65 70 75 80

Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly

85 90 95

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser

100 105 110

Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly

115 120 125

Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro

130 135 140

Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly

145 150 155 160

Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala

165 170 175

Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Tyr Gly

180 185 190

Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser

195 200 205

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly

210 215 220

Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala

225 230 235 240

Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser

245 250 255

Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Tyr Gly Pro Gly Gln

260 265 270

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser

275 280 285

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Pro Ser Ala Ala Ala

290 295 300

Ala Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala

305 310 315 320

Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln

325 330 335

Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly

340 345 350

Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser

355 360 365

Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala

370 375 380

Ala Ala Ala Ala Ala Gly Ser Tyr Gln Gln Gly Pro Gly Gln Gln Gly

385 390 395 400

Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly

405 410 415

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr

420 425 430

Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala

435 440 445

Ala Gly Ser Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro

450 455 460

Gly Gln Ser Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly

465 470 475 480

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro

485 490 495

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala

500 505 510

Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln

515 520 525

Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly

530 535 540

Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln

545 550 555 560

Gly Pro Gly Ala Ser

565

<210> 9

<211> 2364

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT799

<400> 9

Met Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

20 25 30

Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly

35 40 45

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro

50 55 60

Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

65 70 75 80

Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly Gln

85 90 95

Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

100 105 110

Gly Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala

115 120 125

Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr

130 135 140

Gly Pro Gly Ala Ser Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly

145 150 155 160

Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro

165 170 175

Gly Gln Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

180 185 190

Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly

195 200 205

Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala

210 215 220

Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser

225 230 235 240

Ala Ala Ala Ala Ala Gly Gln Tyr Gly Tyr Gly Pro Gly Gln Gln Gly

245 250 255

Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln

260 265 270

Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

275 280 285

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

290 295 300

Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Tyr Gly

305 310 315 320

Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser

325 330 335

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

340 345 350

Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln

355 360 365

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly

370 375 380

Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

385 390 395 400

Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala

405 410 415

Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr

420 425 430

Gly Pro Gly Gln Ser Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro

435 440 445

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro

450 455 460

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

465 470 475 480

Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly

485 490 495

Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser

500 505 510

Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly

515 520 525

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly

530 535 540

Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser

545 550 555 560

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala

565 570 575

Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gln

580 585 590

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln

595 600 605

Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Ser Gly Gln Tyr

610 615 620

Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala

625 630 635 640

Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

645 650 655

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly

660 665 670

Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

675 680 685

Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser

690 695 700

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

705 710 715 720

Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser

725 730 735

Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala

740 745 750

Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro

755 760 765

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly

770 775 780

Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly

785 790 795 800

Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

805 810 815

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

820 825 830

Gly Gln Tyr Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

835 840 845

Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln

850 855 860

Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln

865 870 875 880

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

885 890 895

Gly Pro Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly

900 905 910

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

915 920 925

Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln

930 935 940

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln

945 950 955 960

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr

965 970 975

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly

980 985 990

Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Gln

995 1000 1005

Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln

1010 1015 1020

Ser Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro

1025 1030 1035

Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln

1040 1045 1050

Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly

1055 1060 1065

Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly

1070 1075 1080

Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

1085 1090 1095

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln

1100 1105 1110

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

1115 1120 1125

Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

1130 1135 1140

Gln Ser Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr

1145 1150 1155

Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly

1160 1165 1170

Pro Gly Ala Ser Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser

1175 1180 1185

Ala Ala Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser Gly Gln Gln

1190 1195 1200

Gly Pro Gly Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

1205 1210 1215

Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro

1220 1225 1230

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala

1235 1240 1245

Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser

1250 1255 1260

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro

1265 1270 1275

Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro

1280 1285 1290

Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro

1295 1300 1305

Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser

1310 1315 1320

Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser

1325 1330 1335

Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Tyr

1340 1345 1350

Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser

1355 1360 1365

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser

1370 1375 1380

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala

1385 1390 1395

Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser

1400 1405 1410

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Tyr Gly Pro Gly Gln

1415 1420 1425

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly

1430 1435 1440

Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala

1445 1450 1455

Ala Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

1460 1465 1470

Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln

1475 1480 1485

Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly Gln Gln

1490 1495 1500

Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln

1505 1510 1515

Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala

1520 1525 1530

Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly

1535 1540 1545

Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr

1550 1555 1560

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr

1565 1570 1575

Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala

1580 1585 1590

Gly Gln Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro

1595 1600 1605

Gly Gln Ser Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr

1610 1615 1620

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro

1625 1630 1635

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala

1640 1645 1650

Ala Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln

1655 1660 1665

Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

1670 1675 1680

Gly Gln Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro

1685 1690 1695

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala

1700 1705 1710

Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly

1715 1720 1725

Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly

1730 1735 1740

Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln

1745 1750 1755

Gln Gly Pro Gly Ala Ser Gly Gln Gln Gly Pro Tyr Gly Pro Gly

1760 1765 1770

Ala Ser Ala Ala Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser Gly

1775 1780 1785

Gln Gln Gly Pro Gly Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln

1790 1795 1800

Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

1805 1810 1815

Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser

1820 1825 1830

Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

1835 1840 1845

Ala Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln

1850 1855 1860

Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser

1865 1870 1875

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala

1880 1885 1890

Gly Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly

1895 1900 1905

Ala Ser Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser

1910 1915 1920

Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly

1925 1930 1935

Gln Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

1940 1945 1950

Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser

1955 1960 1965

Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser

1970 1975 1980

Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro

1985 1990 1995

Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Tyr Gly Pro

2000 2005 2010

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly

2015 2020 2025

Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

2030 2035 2040

Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly

2045 2050 2055

Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly

2060 2065 2070

Gln Gln Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly

2075 2080 2085

Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

2090 2095 2100

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln

2105 2110 2115

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln

2120 2125 2130

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly

2135 2140 2145

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

2150 2155 2160

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala

2165 2170 2175

Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr

2180 2185 2190

Gly Pro Gly Gln Ser Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly

2195 2200 2205

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

2210 2215 2220

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala

2225 2230 2235

Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser

2240 2245 2250

Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln

2255 2260 2265

Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln

2270 2275 2280

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

2285 2290 2295

Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro

2300 2305 2310

Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly Pro Gly Gln

2315 2320 2325

Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser

2330 2335 2340

Gly Gln Gln Gly Ser Ser Val Asp Lys Leu Ala Ala Ala Leu Glu

2345 2350 2355

His His His His His His

2360

<210> 10

<211> 597

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT313

<400> 10

Met Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Gly Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

20 25 30

Gly Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Gly Gln Gly

35 40 45

Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro

50 55 60

Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Ser Ala Ser Ala Ala Ala

65 70 75 80

Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Ala Ala

85 90 95

Ala Ala Ala Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Gly Gln Gln

100 105 110

Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Ser Gly

115 120 125

Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro

130 135 140

Gly Ser Gly Gly Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

145 150 155 160

Ala Ala Ala Ala Gly Pro Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro

165 170 175

Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly

180 185 190

Gly Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly

195 200 205

Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gly Ser Ala Ala

210 215 220

Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr

225 230 235 240

Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gly Gln Gly Pro Tyr Gly

245 250 255

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Tyr Gly Pro

260 265 270

Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

275 280 285

Gly Gly Asn Gly Pro Gly Ser Gly Gly Tyr Gly Pro Gly Gln Gln Gly

290 295 300

Pro Gly Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Gly Gln Gly Pro

305 310 315 320

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro

325 330 335

Gly Gly Gln Gly Pro Gly Gly Tyr Gly Pro Gly Ser Ser Ala Ala Ala

340 345 350

Ala Ala Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala

355 360 365

Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly

370 375 380

Pro Gly Gly Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gln Gln Gly Pro

385 390 395 400

Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

405 410 415

Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

420 425 430

Ala Ala Gly Pro Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Ser Ala

435 440 445

Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Ser Gly Pro Gly Gly Tyr

450 455 460

Gly Pro Tyr Gly Pro Gly Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly

465 470 475 480

Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

485 490 495

Ala Ala Ala Gly Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro

500 505 510

Gly Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gly Tyr Gly

515 520 525

Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly Asn Gly Pro Gly Ser

530 535 540

Gly Gly Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gly Ser Ala Ala Ala

545 550 555 560

Ala Ala Gly Gly Tyr Gln Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly

565 570 575

Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln

580 585 590

Gly Pro Gly Ala Ser

595

<210> 11

<211> 12

<212> PRT

<213> Artificial sequence

<220>

<223> His tag

<400> 11

Met His His His His His His Ser Ser Gly Ser Ser

1 5 10

<210> 12

<211> 608

<212> PRT

<213> Artificial sequence

<220>

<223> PRT380

<400> 12

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Gln

1 5 10 15

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln

20 25 30

Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala

35 40 45

Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly

50 55 60

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro

65 70 75 80

Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

85 90 95

Ser Gly Gln Gln Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln

100 105 110

Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser

115 120 125

Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly

130 135 140

Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr

145 150 155 160

Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

165 170 175

Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala

180 185 190

Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Tyr

195 200 205

Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln

210 215 220

Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Ser

225 230 235 240

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala

245 250 255

Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala

260 265 270

Ala Ala Ala Ala Gly Gln Tyr Gly Tyr Gly Pro Gly Gln Gln Gly Pro

275 280 285

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Asn Gly Pro

290 295 300

Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala

305 310 315 320

Ala Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala

325 330 335

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

340 345 350

Gly Gln Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly

355 360 365

Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly

370 375 380

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala

385 390 395 400

Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro

405 410 415

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln

420 425 430

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

435 440 445

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala

450 455 460

Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro

465 470 475 480

Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly

485 490 495

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln

500 505 510

Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala

515 520 525

Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Ala

530 535 540

Ala Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro

545 550 555 560

Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Gln Tyr

565 570 575

Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

580 585 590

Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser

595 600 605

<210> 13

<211> 601

<212> PRT

<213> Artificial sequence

<220>

<223> PRT410

<400> 13

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Gln

1 5 10 15

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln

20 25 30

Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Ser Gly Gln Tyr

35 40 45

Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly

85 90 95

Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

100 105 110

Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser

115 120 125

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly

195 200 205

Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly

210 215 220

Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Gln Tyr Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln

275 280 285

Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

305 310 315 320

Gly Pro Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln

355 360 365

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly

405 410 415

Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Gln

420 425 430

Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser

435 440 445

Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

465 470 475 480

Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln

500 505 510

Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

515 520 525

Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln

545 550 555 560

Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly Pro

565 570 575

Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Gln Gln Gly Pro Gly Ala Ser

595 600

<210> 14

<211> 576

<212> PRT

<213> Artificial sequence

<220>

<223> PRT525

<400> 14

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Gln

1 5 10 15

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala

20 25 30

Gly Ser Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Ser Gly

35 40 45

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser

50 55 60

Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly

65 70 75 80

Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro

85 90 95

Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly

100 105 110

Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala

115 120 125

Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr

130 135 140

Gly Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr

145 150 155 160

Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Tyr Gly

165 170 175

Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala

180 185 190

Gly Ser Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Tyr Ala Ser Ala

195 200 205

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly Gln Gln

210 215 220

Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly Pro Gly

225 230 235 240

Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro

245 250 255

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

260 265 270

Ala Ala Gly Ser Tyr Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly

275 280 285

Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro

290 295 300

Gly Gln Gln Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly

305 310 315 320

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala

325 330 335

Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Tyr

340 345 350

Gly Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

355 360 365

Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln

370 375 380

Gln Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala

385 390 395 400

Gly Ser Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

405 410 415

Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

420 425 430

Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln

435 440 445

Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly

450 455 460

Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser Gly Pro

465 470 475 480

Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

485 490 495

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln Gly Pro

500 505 510

Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

515 520 525

Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser

530 535 540

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Pro Ser Ala Ala Ala

545 550 555 560

Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser

565 570 575

<210> 15

<211> 2375

<212> PRT

<213> Artificial sequence

<220>

<223> PRT799

<400> 15

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Gln

1 5 10 15

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln

20 25 30

Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Ser Gly Gln Tyr

35 40 45

Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly

85 90 95

Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

100 105 110

Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser

115 120 125

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly

195 200 205

Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly

210 215 220

Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Gln Tyr Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln

275 280 285

Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

305 310 315 320

Gly Pro Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln

355 360 365

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly

405 410 415

Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Gln

420 425 430

Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser

435 440 445

Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

465 470 475 480

Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln

500 505 510

Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

515 520 525

Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln

545 550 555 560

Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly Pro

565 570 575

Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gln Gln Gly Pro Tyr Gly

595 600 605

Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser

610 615 620

Gly Gln Gln Gly Pro Gly Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln

625 630 635 640

Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly

645 650 655

Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala

660 665 670

Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly

675 680 685

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser

690 695 700

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Gln

705 710 715 720

Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln

725 730 735

Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Tyr

740 745 750

Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly

755 760 765

Ser Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Tyr Ala Ser Ala Ala

770 775 780

Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr

785 790 795 800

Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly

805 810 815

Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro

820 825 830

Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Tyr

835 840 845

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn

850 855 860

Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

865 870 875 880

Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro

885 890 895

Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln

900 905 910

Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly

915 920 925

Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly

930 935 940

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala

945 950 955 960

Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro

965 970 975

Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser

980 985 990

Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly

995 1000 1005

Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly

1010 1015 1020

Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser Gly Pro Gly

1025 1030 1035

Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

1040 1045 1050

Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Tyr

1055 1060 1065

Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

1070 1075 1080

Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly

1085 1090 1095

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala

1100 1105 1110

Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr

1115 1120 1125

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser

1130 1135 1140

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser

1145 1150 1155

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala

1160 1165 1170

Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser

1175 1180 1185

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala

1190 1195 1200

Ala Gly Gln Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln

1205 1210 1215

Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly

1220 1225 1230

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly

1235 1240 1245

Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly

1250 1255 1260

Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gln Tyr Gly

1265 1270 1275

Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala

1280 1285 1290

Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly

1295 1300 1305

Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln

1310 1315 1320

Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln

1325 1330 1335

Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala

1340 1345 1350

Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Tyr Ala

1355 1360 1365

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln

1370 1375 1380

Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly

1385 1390 1395

Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly

1400 1405 1410

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala

1415 1420 1425

Ala Ala Gly Gln Tyr Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr

1430 1435 1440

Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr

1445 1450 1455

Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

1460 1465 1470

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1475 1480 1485

Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

1490 1495 1500

Tyr Gly Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly

1505 1510 1515

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly

1520 1525 1530

Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

1535 1540 1545

Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro

1550 1555 1560

Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala

1565 1570 1575

Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Gln

1580 1585 1590

Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly

1595 1600 1605

Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser Gly

1610 1615 1620

Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala

1625 1630 1635

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly

1640 1645 1650

Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly

1655 1660 1665

Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly

1670 1675 1680

Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala

1685 1690 1695

Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly

1700 1705 1710

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

1715 1720 1725

Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser

1730 1735 1740

Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser

1745 1750 1755

Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

1760 1765 1770

Ala Ser Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1775 1780 1785

Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro

1790 1795 1800

Gly Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

1805 1810 1815

Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln

1820 1825 1830

Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala

1835 1840 1845

Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gln

1850 1855 1860

Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser

1865 1870 1875

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln

1880 1885 1890

Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser

1895 1900 1905

Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro

1910 1915 1920

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala

1925 1930 1935

Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro

1940 1945 1950

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro

1955 1960 1965

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln

1970 1975 1980

Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala

1985 1990 1995

Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala

2000 2005 2010

Ala Ala Ala Ala Gly Gln Tyr Gly Tyr Gly Pro Gly Gln Gln Gly

2015 2020 2025

Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly

2030 2035 2040

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala

2045 2050 2055

Ala Ala Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser

2060 2065 2070

Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

2075 2080 2085

Gly Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly Pro

2090 2095 2100

Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly

2105 2110 2115

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala

2120 2125 2130

Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr

2135 2140 2145

Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro

2150 2155 2160

Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro

2165 2170 2175

Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Gln

2180 2185 2190

Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln

2195 2200 2205

Ser Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro

2210 2215 2220

Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Gln

2225 2230 2235

Gln Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly

2240 2245 2250

Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly

2255 2260 2265

Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

2270 2275 2280

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly Pro Gly Gln

2285 2290 2295

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

2300 2305 2310

Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly

2315 2320 2325

Gln Ser Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr

2330 2335 2340

Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly

2345 2350 2355

Ser Ser Val Asp Lys Leu Ala Ala Ala Leu Glu His His His His

2360 2365 2370

His His

2375

<210> 16

<211> 608

<212> PRT

<213> Artificial sequence

<220>

<223> PRT313

<400> 16

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Gly

1 5 10 15

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly

20 25 30

Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gly Ser Ala Ala Ala

35 40 45

Ala Ala Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Gly Gln Gln Gly

50 55 60

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Gly Tyr Gly Pro

65 70 75 80

Gly Gly Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

85 90 95

Ser Gly Gln Gln Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly

100 105 110

Tyr Gly Pro Gly Gly Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser

115 120 125

Ala Ala Ala Ala Ala Gly Gly Tyr Gly Ser Gly Pro Gly Gln Gln Gly

130 135 140

Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gly Tyr

145 150 155 160

Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

165 170 175

Pro Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Ser Ala Ser Ala Ala

180 185 190

Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gly Tyr Gly Pro Tyr

195 200 205

Ala Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Ser Gly Pro Gly Gln

210 215 220

Gln Gly Pro Tyr Gly Pro Gly Gly Ser Ala Ala Ala Ala Ala Gly Ser

225 230 235 240

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala

245 250 255

Ala Ala Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala

260 265 270

Ala Ala Ala Ala Gly Gly Tyr Gly Tyr Gly Pro Gly Gly Gln Gly Pro

275 280 285

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly Asn Gly Pro

290 295 300

Gly Ser Gly Gly Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gly Ser Ala

305 310 315 320

Ala Ala Ala Ala Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala

325 330 335

Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro

340 345 350

Gly Gly Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly

355 360 365

Gly Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly

370 375 380

Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gly Ser Ala

385 390 395 400

Ala Ala Ala Ala Gly Gly Tyr Gln Gln Gly Pro Gly Gly Gln Gly Pro

405 410 415

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gly Gln

420 425 430

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

435 440 445

Gly Tyr Gly Pro Gly Gly Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala

450 455 460

Ala Gly Gly Tyr Gly Ser Gly Pro Gly Gly Tyr Gly Pro Tyr Gly Pro

465 470 475 480

Gly Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly

485 490 495

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly

500 505 510

Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gly Ser Ala Ala

515 520 525

Ala Ala Ala Gly Pro Gly Ser Gly Gly Tyr Gly Pro Gly Ala Ser Ala

530 535 540

Ala Ala Ala Ala Gly Gly Asn Gly Pro Gly Ser Gly Gly Tyr Gly Pro

545 550 555 560

Gly Gln Gln Gly Pro Gly Gly Ser Ala Ala Ala Ala Ala Gly Gly Tyr

565 570 575

Gln Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

580 585 590

Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser

595 600 605

<210> 17

<211> 590

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT399

<400> 17

Met Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Gly Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

20 25 30

Gly Ser Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Gly Gln Gln Gly

35 40 45

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Gly Tyr Gly Pro

50 55 60

Gly Gly Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

65 70 75 80

Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gly Tyr Gly Pro Gly Gly

85 90 95

Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

100 105 110

Gly Gly Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala

115 120 125

Ala Ala Ala Ala Gly Pro Gly Ser Gly Gly Tyr Gly Gln Gly Pro Tyr

130 135 140

Gly Pro Gly Ala Ser Gly Pro Gly Gly Tyr Gly Pro Gly Gly Gln Gly

145 150 155 160

Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro

165 170 175

Gly Gly Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gly Tyr

180 185 190

Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gly Ser Gly

195 200 205

Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala

210 215 220

Ala Ala Ala Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ser Ser

225 230 235 240

Ala Ala Ala Ala Ala Gly Gly Tyr Gly Tyr Gly Pro Gly Gly Gln Gly

245 250 255

Pro Tyr Gly Pro Gly Ala Ser Gly Gly Asn Gly Pro Gly Ser Gly Gly

260 265 270

Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gly Ser Ala Ala Ala Ala Ala

275 280 285

Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

290 295 300

Ala Ala Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Gly Gly Tyr Gly

305 310 315 320

Pro Gly Ser Ser Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ser

325 330 335

Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Gln Gln Gly Pro

340 345 350

Tyr Gly Pro Gly Gly Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gln Gln

355 360 365

Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly

370 375 380

Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

385 390 395 400

Pro Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Ser Ala Ser Ala Ala

405 410 415

Ala Ala Ala Gly Gly Tyr Gly Ser Gly Pro Gly Gly Tyr Gly Pro Tyr

420 425 430

Gly Pro Gly Gly Ser Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro

435 440 445

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro

450 455 460

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gly Ser Ala Ala Ala Ala Ala

465 470 475 480

Gly Pro Gly Ser Gly Gly Tyr Gly Pro Gly Ala Ser Gly Gly Asn Gly

485 490 495

Pro Gly Ser Gly Gly Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gly Ser

500 505 510

Ala Ala Ala Ala Ala Gly Gly Tyr Gln Gln Gly Pro Gly Gly Gln Gly

515 520 525

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly

530 535 540

Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gly Ser Gly Ser

545 550 555 560

Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala

565 570 575

Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser

580 585 590

<210> 18

<211> 601

<212> PRT

<213> Artificial sequence

<220>

<223> PRT399

<400> 18

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Gly

1 5 10 15

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly

20 25 30

Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gly Ser Gly Gly Tyr

35 40 45

Gly Pro Gly Gly Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly

85 90 95

Pro Gly Ala Ser Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Gly Gln

100 105 110

Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Ser

115 120 125

Gly Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Gly Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Gly Tyr Gly Pro Gly Gly Gln Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Gln Gln Gly Pro Gly Gly Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Ser Gly Pro Gly

195 200 205

Gln Gln Gly Pro Tyr Gly Pro Gly Gly Ser Gly Ser Gly Gln Gln Gly

210 215 220

Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Gly Tyr Gly Tyr Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Gly Asn Gly Pro Gly Ser Gly Gly Tyr Gly Pro Gly Gln

275 280 285

Gln Gly Pro Gly Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Gly Gln

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gly Tyr

305 310 315 320

Gly Pro Gly Gly Gln Gly Pro Gly Gly Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gly

355 360 365

Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gln Gln Gly Pro Gly Gly Gln

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gly Gln Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gly Tyr Gly

405 410 415

Pro Gly Gly Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Gly

420 425 430

Tyr Gly Ser Gly Pro Gly Gly Tyr Gly Pro Tyr Gly Pro Gly Gly Ser

435 440 445

Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Pro Gly Gln Gln Gly Pro

465 470 475 480

Tyr Gly Pro Gly Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Gly Tyr Gly Pro Gly Ala Ser Gly Gly Asn Gly Pro Gly Ser Gly Gly

500 505 510

Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gly Ser Ala Ala Ala Ala Ala

515 520 525

Gly Gly Tyr Gln Gln Gly Pro Gly Gly Gln Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Gly Tyr Gly Ser Gly Pro Gly Gln

545 550 555 560

Gln Gly Pro Tyr Gly Pro Gly Gly Ser Gly Ser Gly Gln Gln Gly Pro

565 570 575

Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Gln Gln Gly Pro Gly Ala Ser

595 600

<210> 19

<211> 612

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT720

<400> 19

Met Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

20 25 30

Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly

35 40 45

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Val Leu

50 55 60

Ile Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Ser Ala Ser Ala Ala

65 70 75 80

Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly

85 90 95

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser

100 105 110

Ser Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Val Leu Ile Gly Pro

115 120 125

Gly Gln Gln Val Leu Ile Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala

130 135 140

Gly Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala

145 150 155 160

Ser Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser

165 170 175

Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Val Leu Ile Gly Pro Gly

180 185 190

Gln Tyr Val Leu Ile Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly

195 200 205

Gln Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln

210 215 220

Ser Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser

225 230 235 240

Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr

245 250 255

Val Leu Ile Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr

260 265 270

Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly

275 280 285

Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro

290 295 300

Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Val Leu Ile

305 310 315 320

Gly Pro Tyr Val Leu Ile Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

325 330 335

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Gly

340 345 350

Ser Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala

355 360 365

Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln Val Leu Ile Gly

370 375 380

Pro Tyr Val Leu Ile Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Gly

385 390 395 400

Gln Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala

405 410 415

Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

420 425 430

Ala Ala Ala Gly Pro Gly Gln Tyr Val Leu Ile Gly Pro Gly Gln Gln

435 440 445

Val Leu Ile Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

450 455 460

Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser Gly

465 470 475 480

Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser

485 490 495

Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln Val Leu Ile

500 505 510

Gly Pro Tyr Val Leu Ile Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala

515 520 525

Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly

530 535 540

Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser

545 550 555 560

Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Val Leu Ile Gly Pro Gly

565 570 575

Gln Gln Gly Pro Tyr Val Leu Ile Gly Pro Gly Ala Ser Ala Ala Ala

580 585 590

Ala Ala Gly Pro Gly Ser Gly Gln Gln Val Leu Ile Gly Pro Gly Ala

595 600 605

Ser Val Leu Ile

610

<210> 20

<211> 592

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT665

<400> 20

Met Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Ala Ala Gly Ser Asn Gly Pro Gly Ser Gly Gln Gln Gly

20 25 30

Pro Gly Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

35 40 45

Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

50 55 60

Gln Tyr Val Leu Ile Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala

65 70 75 80

Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly

85 90 95

Ala Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly

100 105 110

Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser

115 120 125

Val Leu Ile Gly Pro Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala

130 135 140

Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr

145 150 155 160

Gly Pro Gly Ala Ser Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly

165 170 175

Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln

180 185 190

Val Leu Ile Gly Pro Gly Gln Tyr Gly Pro Tyr Ala Ser Ala Ala Ala

195 200 205

Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro

210 215 220

Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln

225 230 235 240

Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln

245 250 255

Gln Val Leu Ile Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

260 265 270

Ala Ala Ala Gly Ser Tyr Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr

275 280 285

Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly

290 295 300

Pro Gly Gln Gln Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala

305 310 315 320

Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Gly Pro Gly Ala Ser

325 330 335

Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln Gly

340 345 350

Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly Pro

355 360 365

Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr

370 375 380

Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Gly Pro Gly Pro Ser

385 390 395 400

Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gln Gln Gly Pro Gly Gln

405 410 415

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro

420 425 430

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

435 440 445

Gln Tyr Val Leu Ile Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala

450 455 460

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly Gln Tyr

465 470 475 480

Gly Pro Tyr Gly Pro Gly Gln Ser Gly Pro Gly Ser Gly Gln Gln Gly

485 490 495

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala

500 505 510

Gly Ser Tyr Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Gly Pro

515 520 525

Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln

530 535 540

Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr

545 550 555 560

Gly Pro Gly Gln Gln Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala

565 570 575

Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Val Leu Ile

580 585 590

<210> 21

<211> 619

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT666

<400> 21

Met Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Ala Ala Gly Ser Asn Gly Pro Gly Ser Gly Gln Gln Gly

20 25 30

Pro Gly Gln Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln

35 40 45

Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

50 55 60

Gln Tyr Val Leu Ile Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Ser

65 70 75 80

Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln

85 90 95

Gly Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly

100 105 110

Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser

115 120 125

Tyr Gly Ser Val Leu Ile Gly Pro Gly Gln Gln Val Leu Ile Gly Pro

130 135 140

Tyr Gly Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln

145 150 155 160

Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Tyr

165 170 175

Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala

180 185 190

Ala Gly Ser Gly Gln Gln Val Leu Ile Gly Pro Gly Gln Tyr Val Leu

195 200 205

Ile Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr

210 215 220

Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly

225 230 235 240

Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala

245 250 255

Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr

260 265 270

Val Leu Ile Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly

275 280 285

Ser Tyr Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala

290 295 300

Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln

305 310 315 320

Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln

325 330 335

Gln Val Leu Ile Gly Pro Tyr Val Leu Ile Gly Pro Gly Ala Ser Ala

340 345 350

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln Gly Pro

355 360 365

Gly Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly Pro Tyr

370 375 380

Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly

385 390 395 400

Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Val Leu Ile Gly Pro Gly

405 410 415

Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gln Gln Gly Pro

420 425 430

Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln

435 440 445

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly

450 455 460

Pro Gly Gln Tyr Val Leu Ile Gly Pro Gly Gln Gln Val Leu Ile Gly

465 470 475 480

Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser

485 490 495

Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser Gly Pro Gly

500 505 510

Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

515 520 525

Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln Val Leu Ile

530 535 540

Gly Pro Tyr Val Leu Ile Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala

545 550 555 560

Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln

565 570 575

Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly

580 585 590

Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln

595 600 605

Val Leu Ile Gly Pro Gly Ala Ser Val Leu Ile

610 615

<210> 22

<211> 623

<212> PRT

<213> Artificial sequence

<220>

<223> PRT720

<400> 22

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Gln

1 5 10 15

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln

20 25 30

Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Ser Gly Gln Tyr

35 40 45

Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Gln Tyr Val Leu Ile Gly Pro Gly Gln

65 70 75 80

Gln Val Leu Ile Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro

85 90 95

Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly

100 105 110

Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala Ala Ala Ala

115 120 125

Ala Gly Ser Tyr Gly Ser Val Leu Ile Gly Pro Gly Gln Gln Val Leu

130 135 140

Ile Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

145 150 155 160

Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln

165 170 175

Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala

180 185 190

Gly Ser Gly Gln Gln Val Leu Ile Gly Pro Gly Gln Tyr Val Leu Ile

195 200 205

Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly

210 215 220

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln

225 230 235 240

Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala

245 250 255

Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Val Leu Ile Gly Pro

260 265 270

Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Tyr Gly Pro Gly

275 280 285

Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly

290 295 300

Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala

305 310 315 320

Ala Ala Ala Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Val Leu

325 330 335

Ile Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro

340 345 350

Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly

355 360 365

Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly

370 375 380

Ser Tyr Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Val Leu Ile

385 390 395 400

Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gln Gln Gly

405 410 415

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln

420 425 430

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro

435 440 445

Gly Gln Tyr Val Leu Ile Gly Pro Gly Gln Gln Val Leu Ile Gly Pro

450 455 460

Ser Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly

465 470 475 480

Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser Gly Pro Gly Ser Gly Gln

485 490 495

Gln Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

500 505 510

Gly Ser Tyr Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Val Leu

515 520 525

Ile Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

530 535 540

Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln

545 550 555 560

Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

565 570 575

Gly Gln Tyr Gln Gln Val Leu Ile Gly Pro Gly Gln Gln Gly Pro Tyr

580 585 590

Val Leu Ile Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

595 600 605

Ser Gly Gln Gln Val Leu Ile Gly Pro Gly Ala Ser Val Leu Ile

610 615 620

<210> 23

<211> 603

<212> PRT

<213> Artificial sequence

<220>

<223> PRT665

<400> 23

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Gln

1 5 10 15

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala

20 25 30

Gly Ser Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Ser Gly

35 40 45

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser

50 55 60

Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Val Leu Ile

65 70 75 80

Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala

85 90 95

Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser Gly Gln Tyr

100 105 110

Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser Ser Ala

115 120 125

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Val Leu Ile Gly Pro

130 135 140

Gly Gln Gln Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Ala Ala Gly

145 150 155 160

Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser

165 170 175

Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala

180 185 190

Ala Ala Ala Ala Ala Ala Gly Ser Gly Gln Gln Val Leu Ile Gly Pro

195 200 205

Gly Gln Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly

210 215 220

Ser Tyr Gly Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Gln

225 230 235 240

Ser Gly Ser Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Ala Ser

245 250 255

Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Val Leu Ile Gly

260 265 270

Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser

275 280 285

Tyr Gly Tyr Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser

290 295 300

Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly

305 310 315 320

Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln

325 330 335

Val Leu Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

340 345 350

Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Tyr Gly

355 360 365

Pro Gly Ser Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser

370 375 380

Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln

385 390 395 400

Val Leu Ile Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala

405 410 415

Ala Ala Gly Ser Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro Tyr Gly

420 425 430

Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala

435 440 445

Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Val Leu Ile

450 455 460

Gly Pro Gly Gln Gln Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala

465 470 475 480

Ala Gly Ser Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro

485 490 495

Gly Gln Ser Gly Pro Gly Ser Gly Gln Gln Gly Gln Gly Pro Tyr Gly

500 505 510

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro

515 520 525

Gly Gln Gln Val Leu Ile Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala

530 535 540

Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala

545 550 555 560

Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln

565 570 575

Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser

580 585 590

Gly Gln Gln Gly Pro Gly Ala Ser Val Leu Ile

595 600

<210> 24

<211> 630

<212> PRT

<213> Artificial sequence

<220>

<223> PRT666

<400> 24

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Gln

1 5 10 15

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala

20 25 30

Gly Ser Asn Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Gln Ser Gly

35 40 45

Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly Ser

50 55 60

Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Val Leu Ile

65 70 75 80

Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Ser Ala Ser Ala Ala Ala

85 90 95

Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Gly Pro Gly Ala Ser

100 105 110

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Gln Gly Pro Gly

115 120 125

Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Val Leu

130 135 140

Ile Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Gly Ser Ala Ala

145 150 155 160

Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro

165 170 175

Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Tyr Gly Pro Gly Gln Gln

180 185 190

Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Gly Gln

195 200 205

Gln Val Leu Ile Gly Pro Gly Gln Tyr Val Leu Ile Gly Pro Tyr Ala

210 215 220

Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly

225 230 235 240

Gln Gln Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Gln Gln Gly

245 250 255

Pro Gly Gln Gln Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala

260 265 270

Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Val Leu Ile Gly Pro

275 280 285

Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Tyr Gly

290 295 300

Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly

305 310 315 320

Pro Gly Ser Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Pro Ser

325 330 335

Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Gln Val Leu Ile Gly

340 345 350

Pro Tyr Val Leu Ile Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala

355 360 365

Ala Gly Ser Tyr Gly Pro Gly Gln Gln Gly Pro Gly Gln Tyr Gly Pro

370 375 380

Gly Ser Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro Gly Ser Ser

385 390 395 400

Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Gln Gln Val

405 410 415

Leu Ile Gly Pro Tyr Val Leu Ile Gly Pro Gly Pro Ser Ala Ala Ala

420 425 430

Ala Ala Ala Ala Gly Ser Tyr Gln Gln Gly Pro Gly Gln Gln Gly Pro

435 440 445

Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Gln Gly Pro Tyr Gly Pro

450 455 460

Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Val

465 470 475 480

Leu Ile Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Ser Ala Ser Ala

485 490 495

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly Gln Tyr

500 505 510

Gly Pro Tyr Gly Pro Gly Gln Ser Gly Pro Gly Ser Gly Gln Gln Gly

515 520 525

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala

530 535 540

Gly Ser Tyr Gly Pro Gly Gln Gln Val Leu Ile Gly Pro Tyr Val Leu

545 550 555 560

Ile Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

565 570 575

Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser

580 585 590

Gly Gln Tyr Gly Pro Gly Gln Gln Gly Pro Gly Pro Ser Ala Ala Ala

595 600 605

Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Gln Val Leu Ile Gly Pro

610 615 620

Gly Ala Ser Val Leu Ile

625 630

<210> 25

<211> 593

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT888

<400> 25

Met Gly Ser Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala

1 5 10 15

Ser Ala Ala Ala Ala Ala Gly Gln Asn Gly Pro Gly Ser Gly Val Leu

20 25 30

Gly Pro Gly Gln Ser Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly

35 40 45

Val Leu Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln

50 55 60

Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala

65 70 75 80

Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ala Ser Gly Gln Tyr Gly

85 90 95

Pro Gly Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser Ser Ala Ala

100 105 110

Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Val Leu Gly Pro Tyr

115 120 125

Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Gln

130 135 140

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Tyr Gly Pro Gly

145 150 155 160

Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Val

165 170 175

Leu Gly Pro Gly Gln Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala

180 185 190

Gly Gln Tyr Gly Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

195 200 205

Gln Ser Gly Ser Gly Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Ala

210 215 220

Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro

225 230 235 240

Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Tyr Gly Pro Gly

245 250 255

Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly

260 265 270

Ser Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly Gln Ser Ala Ala

275 280 285

Ala Ala Ala Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser

290 295 300

Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly

305 310 315 320

Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly

325 330 335

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Val

340 345 350

Leu Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Gln

355 360 365

Tyr Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser

370 375 380

Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

385 390 395 400

Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala

405 410 415

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Gln Tyr

420 425 430

Gly Pro Tyr Gly Pro Gly Gln Ser Gly Pro Gly Ser Gly Val Leu Gly

435 440 445

Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln

450 455 460

Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Gln Ser Ala Ala

465 470 475 480

Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly

485 490 495

Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro

500 505 510

Gly Gln Ser Ala Ala Ala Ala Ala Gly Gln Tyr Val Leu Gly Pro Gly

515 520 525

Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

530 535 540

Gln Tyr Gly Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Gln

545 550 555 560

Ser Gly Ser Gly Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Ala Ser

565 570 575

Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ala

580 585 590

Ser

<210> 26

<211> 590

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT965

<400> 26

Met Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Ala Asn Gly Pro Gly Ser Gly Thr Ser Gly Pro Gly

20 25 30

Ala Ser Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro Gly Thr Ser Gly

35 40 45

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Ala Tyr Gly Pro

50 55 60

Gly Thr Ser Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

65 70 75 80

Ser Gly Thr Ser Gly Pro Gly Ala Ser Gly Ala Tyr Gly Pro Gly Thr

85 90 95

Ser Gly Pro Gly Thr Ser Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

100 105 110

Gly Ala Tyr Gly Ser Gly Pro Gly Thr Ser Gly Pro Tyr Gly Ser Ala

115 120 125

Ala Ala Ala Ala Gly Pro Gly Ser Gly Ala Tyr Gly Ala Gly Pro Tyr

130 135 140

Gly Pro Gly Ala Ser Gly Pro Gly Ala Tyr Gly Pro Gly Thr Ser Gly

145 150 155 160

Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Thr Ser Gly Pro

165 170 175

Gly Ala Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Ala Tyr

180 185 190

Gly Ser Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ala Ser Gly

195 200 205

Ser Gly Thr Ser Gly Pro Gly Thr Ser Gly Pro Tyr Ala Ser Ala Ala

210 215 220

Ala Ala Ala Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ser Ser

225 230 235 240

Ala Ala Ala Ala Ala Gly Ala Tyr Gly Tyr Gly Pro Gly Thr Ser Gly

245 250 255

Pro Tyr Gly Pro Gly Ala Ser Gly Ala Asn Gly Pro Gly Ser Gly Ala

260 265 270

Tyr Gly Pro Gly Thr Ser Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

275 280 285

Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

290 295 300

Ala Ala Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro Gly Ala Tyr Gly

305 310 315 320

Pro Gly Ser Ser Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ser

325 330 335

Ser Ala Ala Ala Ala Ala Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro

340 345 350

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ala Tyr Thr Ser

355 360 365

Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly

370 375 380

Thr Ser Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

385 390 395 400

Pro Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro Ser Ala Ser Ala Ala

405 410 415

Ala Ala Ala Gly Ala Tyr Gly Ser Gly Pro Gly Ala Tyr Gly Pro Tyr

420 425 430

Gly Pro Gly Ala Ser Gly Pro Gly Ser Gly Thr Ser Gly Ala Gly Pro

435 440 445

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ala Tyr Gly Pro

450 455 460

Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

465 470 475 480

Gly Pro Gly Ser Gly Ala Tyr Gly Pro Gly Ala Ser Gly Ala Asn Gly

485 490 495

Pro Gly Ser Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro Gly Ala Ser

500 505 510

Ala Ala Ala Ala Ala Gly Ala Tyr Thr Ser Gly Pro Gly Thr Ser Gly

515 520 525

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ala Tyr Gly

530 535 540

Ser Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ala Ser Gly Ser

545 550 555 560

Gly Thr Ser Gly Pro Gly Thr Ser Gly Pro Tyr Ala Ser Ala Ala Ala

565 570 575

Ala Ala Gly Pro Gly Ser Gly Thr Ser Gly Pro Gly Ala Ser

580 585 590

<210> 27

<211> 593

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT889

<400> 27

Met Gly Ser Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala

1 5 10 15

Ser Ala Ala Ala Ala Ala Gly Ile Asn Gly Pro Gly Ser Gly Val Leu

20 25 30

Gly Pro Gly Ile Ser Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly

35 40 45

Val Leu Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile

50 55 60

Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala

65 70 75 80

Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ala Ser Gly Ile Tyr Gly

85 90 95

Pro Gly Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser Ser Ala Ala

100 105 110

Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly Val Leu Gly Pro Tyr

115 120 125

Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Ile Tyr Gly Ile

130 135 140

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Ile Tyr Gly Pro Gly

145 150 155 160

Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Val

165 170 175

Leu Gly Pro Gly Ile Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala

180 185 190

Gly Ile Tyr Gly Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

195 200 205

Ile Ser Gly Ser Gly Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Ala

210 215 220

Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro

225 230 235 240

Gly Ser Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Tyr Gly Pro Gly

245 250 255

Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly

260 265 270

Ser Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly Ile Ser Ala Ala

275 280 285

Ala Ala Ala Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser

290 295 300

Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly

305 310 315 320

Ile Tyr Gly Pro Gly Ser Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly

325 330 335

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro Gly Val

340 345 350

Leu Gly Pro Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Ile

355 360 365

Tyr Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser

370 375 380

Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

385 390 395 400

Ala Ala Gly Pro Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala

405 410 415

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly Ile Tyr

420 425 430

Gly Pro Tyr Gly Pro Gly Ile Ser Gly Pro Gly Ser Gly Val Leu Gly

435 440 445

Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile

450 455 460

Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ile Ser Ala Ala

465 470 475 480

Ala Ala Ala Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Ala Ser Gly

485 490 495

Ile Asn Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro

500 505 510

Gly Ile Ser Ala Ala Ala Ala Ala Gly Ile Tyr Val Leu Gly Pro Gly

515 520 525

Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

530 535 540

Ile Tyr Gly Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ile

545 550 555 560

Ser Gly Ser Gly Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Ala Ser

565 570 575

Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ala

580 585 590

Ser

<210> 28

<211> 590

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT916

<400> 28

Met Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Leu Asn Gly Pro Gly Ser Gly Val Ile Gly Pro Gly

20 25 30

Leu Ser Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro Gly Val Ile Gly

35 40 45

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Leu Tyr Gly Pro

50 55 60

Gly Val Ile Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

65 70 75 80

Ser Gly Val Ile Gly Pro Gly Ala Ser Gly Leu Tyr Gly Pro Gly Val

85 90 95

Ile Gly Pro Gly Val Ile Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

100 105 110

Gly Leu Tyr Gly Ser Gly Pro Gly Val Ile Gly Pro Tyr Gly Ser Ala

115 120 125

Ala Ala Ala Ala Gly Pro Gly Ser Gly Leu Tyr Gly Leu Gly Pro Tyr

130 135 140

Gly Pro Gly Ala Ser Gly Pro Gly Leu Tyr Gly Pro Gly Val Ile Gly

145 150 155 160

Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Val Ile Gly Pro

165 170 175

Gly Leu Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Leu Tyr

180 185 190

Gly Ser Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly Leu Ser Gly

195 200 205

Ser Gly Val Ile Gly Pro Gly Val Ile Gly Pro Tyr Ala Ser Ala Ala

210 215 220

Ala Ala Ala Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly Ser Ser

225 230 235 240

Ala Ala Ala Ala Ala Gly Leu Tyr Gly Tyr Gly Pro Gly Val Ile Gly

245 250 255

Pro Tyr Gly Pro Gly Ala Ser Gly Leu Asn Gly Pro Gly Ser Gly Leu

260 265 270

Tyr Gly Pro Gly Val Ile Gly Pro Gly Leu Ser Ala Ala Ala Ala Ala

275 280 285

Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

290 295 300

Ala Ala Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro Gly Leu Tyr Gly

305 310 315 320

Pro Gly Ser Ser Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly Ser

325 330 335

Ser Ala Ala Ala Ala Ala Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro

340 345 350

Tyr Gly Pro Gly Leu Ser Ala Ala Ala Ala Ala Gly Leu Tyr Val Ile

355 360 365

Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly

370 375 380

Val Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

385 390 395 400

Pro Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro Ser Ala Ser Ala Ala

405 410 415

Ala Ala Ala Gly Leu Tyr Gly Ser Gly Pro Gly Leu Tyr Gly Pro Tyr

420 425 430

Gly Pro Gly Leu Ser Gly Pro Gly Ser Gly Val Ile Gly Leu Gly Pro

435 440 445

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Leu Tyr Gly Pro

450 455 460

Gly Val Ile Gly Pro Tyr Gly Pro Gly Leu Ser Ala Ala Ala Ala Ala

465 470 475 480

Gly Pro Gly Ser Gly Leu Tyr Gly Pro Gly Ala Ser Gly Leu Asn Gly

485 490 495

Pro Gly Ser Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro Gly Leu Ser

500 505 510

Ala Ala Ala Ala Ala Gly Leu Tyr Val Ile Gly Pro Gly Val Ile Gly

515 520 525

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Leu Tyr Gly

530 535 540

Ser Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly Leu Ser Gly Ser

545 550 555 560

Gly Val Ile Gly Pro Gly Val Ile Gly Pro Tyr Ala Ser Ala Ala Ala

565 570 575

Ala Ala Gly Pro Gly Ser Gly Val Ile Gly Pro Gly Ala Ser

580 585 590

<210> 29

<211> 590

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT918

<400> 29

Met Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Ile Asn Gly Pro Gly Ser Gly Val Phe Gly Pro Gly

20 25 30

Ile Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Val Phe Gly

35 40 45

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly Pro

50 55 60

Gly Val Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

65 70 75 80

Ser Gly Val Phe Gly Pro Gly Ala Ser Gly Ile Tyr Gly Pro Gly Val

85 90 95

Phe Gly Pro Gly Val Phe Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

100 105 110

Gly Ile Tyr Gly Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Ser Ala

115 120 125

Ala Ala Ala Ala Gly Pro Gly Ser Gly Ile Tyr Gly Ile Gly Pro Tyr

130 135 140

Gly Pro Gly Ala Ser Gly Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly

145 150 155 160

Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Val Phe Gly Pro

165 170 175

Gly Ile Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr

180 185 190

Gly Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Gly

195 200 205

Ser Gly Val Phe Gly Pro Gly Val Phe Gly Pro Tyr Ala Ser Ala Ala

210 215 220

Ala Ala Ala Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser Ser

225 230 235 240

Ala Ala Ala Ala Ala Gly Ile Tyr Gly Tyr Gly Pro Gly Val Phe Gly

245 250 255

Pro Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile

260 265 270

Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala

275 280 285

Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

290 295 300

Ala Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Tyr Gly

305 310 315 320

Pro Gly Ser Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser

325 330 335

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro

340 345 350

Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Ile Tyr Val Phe

355 360 365

Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly

370 375 380

Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

385 390 395 400

Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Ser Ala Ser Ala Ala

405 410 415

Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly Ile Tyr Gly Pro Tyr

420 425 430

Gly Pro Gly Ile Ser Gly Pro Gly Ser Gly Val Phe Gly Ile Gly Pro

435 440 445

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro

450 455 460

Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala

465 470 475 480

Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly

485 490 495

Pro Gly Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Ser

500 505 510

Ala Ala Ala Ala Ala Gly Ile Tyr Val Phe Gly Pro Gly Val Phe Gly

515 520 525

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly

530 535 540

Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser

545 550 555 560

Gly Val Phe Gly Pro Gly Val Phe Gly Pro Tyr Ala Ser Ala Ala Ala

565 570 575

Ala Ala Gly Pro Gly Ser Gly Val Phe Gly Pro Gly Ala Ser

580 585 590

<210> 30

<211> 565

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT699

<400> 30

Met Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Ala Ala Gly Ser Asn Gly Pro Gly Ser Gly Val Leu Gly

20 25 30

Pro Gly Gln Ser Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly Val

35 40 45

Leu Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

50 55 60

Gln Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala

65 70 75 80

Ala Ala Ala Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ala Ser Gly

85 90 95

Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser

100 105 110

Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly

115 120 125

Val Leu Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro

130 135 140

Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly

145 150 155 160

Pro Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala

165 170 175

Ala Ala Ala Ala Ala Gly Ser Gly Val Leu Gly Pro Gly Gln Tyr Gly

180 185 190

Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser

195 200 205

Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly

210 215 220

Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Ala Ser Ala Ala Ala Ala

225 230 235 240

Ala Ala Ala Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ser Ser

245 250 255

Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Tyr Gly Pro Gly Val

260 265 270

Leu Gly Pro Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser

275 280 285

Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly Pro Ser Ala Ala Ala

290 295 300

Ala Ala Ala Ala Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala

305 310 315 320

Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Val Leu

325 330 335

Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly Pro Gly Val Leu Gly

340 345 350

Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser

355 360 365

Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala

370 375 380

Ala Ala Ala Ala Ala Gly Ser Tyr Val Leu Gly Pro Gly Val Leu Gly

385 390 395 400

Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly

405 410 415

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr

420 425 430

Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala

435 440 445

Ala Gly Ser Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro

450 455 460

Gly Gln Ser Gly Pro Gly Ser Gly Val Leu Gly Gln Gly Pro Tyr Gly

465 470 475 480

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro

485 490 495

Gly Val Leu Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala

500 505 510

Ala Ala Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln

515 520 525

Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly

530 535 540

Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Leu

545 550 555 560

Gly Pro Gly Ala Ser

565

<210> 31

<211> 565

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT698

<400> 31

Met Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Ala Ala Gly Ser Asn Gly Pro Gly Ser Gly Val Leu Gly

20 25 30

Pro Gly Ile Ser Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly Val

35 40 45

Leu Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

50 55 60

Ile Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala

65 70 75 80

Ala Ala Ala Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ala Ser Gly

85 90 95

Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser

100 105 110

Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly

115 120 125

Val Leu Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro

130 135 140

Gly Ser Gly Ile Tyr Gly Ile Gly Pro Tyr Gly Pro Gly Ala Ser Gly

145 150 155 160

Pro Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala

165 170 175

Ala Ala Ala Ala Ala Gly Ser Gly Val Leu Gly Pro Gly Ile Tyr Gly

180 185 190

Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser

195 200 205

Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly

210 215 220

Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Ala Ser Ala Ala Ala Ala

225 230 235 240

Ala Ala Ala Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ser Ser

245 250 255

Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Tyr Gly Pro Gly Val

260 265 270

Leu Gly Pro Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly Ser

275 280 285

Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly Pro Ser Ala Ala Ala

290 295 300

Ala Ala Ala Ala Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala

305 310 315 320

Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Val Leu

325 330 335

Gly Pro Gly Ile Tyr Gly Pro Gly Ser Ser Gly Pro Gly Val Leu Gly

340 345 350

Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser

355 360 365

Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala

370 375 380

Ala Ala Ala Ala Ala Gly Ser Tyr Val Leu Gly Pro Gly Val Leu Gly

385 390 395 400

Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly

405 410 415

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr

420 425 430

Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala

435 440 445

Ala Gly Ser Tyr Gly Ser Gly Pro Gly Ile Tyr Gly Pro Tyr Gly Pro

450 455 460

Gly Ile Ser Gly Pro Gly Ser Gly Val Leu Gly Ile Gly Pro Tyr Gly

465 470 475 480

Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro

485 490 495

Gly Val Leu Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala

500 505 510

Ala Ala Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Ala Ser Gly Ile

515 520 525

Asn Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly

530 535 540

Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Leu

545 550 555 560

Gly Pro Gly Ala Ser

565

<210> 32

<211> 1179

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT966

<400> 32

Met Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Ile Asn Gly Pro Gly Ser Gly Val Phe Gly Pro Gly

20 25 30

Ile Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Val Phe Gly

35 40 45

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly Pro

50 55 60

Gly Val Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

65 70 75 80

Ser Gly Val Phe Gly Pro Gly Ala Ser Gly Ile Tyr Gly Pro Gly Val

85 90 95

Phe Gly Pro Gly Val Phe Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

100 105 110

Gly Ile Tyr Gly Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Ser Ala

115 120 125

Ala Ala Ala Ala Gly Pro Gly Ser Gly Ile Tyr Gly Ile Gly Pro Tyr

130 135 140

Gly Pro Gly Ala Ser Gly Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly

145 150 155 160

Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Val Phe Gly Pro

165 170 175

Gly Ile Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr

180 185 190

Gly Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Gly

195 200 205

Ser Gly Val Phe Gly Pro Gly Val Phe Gly Pro Tyr Ala Ser Ala Ala

210 215 220

Ala Ala Ala Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser Ser

225 230 235 240

Ala Ala Ala Ala Ala Gly Ile Tyr Gly Tyr Gly Pro Gly Val Phe Gly

245 250 255

Pro Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile

260 265 270

Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala

275 280 285

Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

290 295 300

Ala Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Tyr Gly

305 310 315 320

Pro Gly Ser Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser

325 330 335

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro

340 345 350

Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Ile Tyr Val Phe

355 360 365

Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly

370 375 380

Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

385 390 395 400

Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Ser Ala Ser Ala Ala

405 410 415

Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly Ile Tyr Gly Pro Tyr

420 425 430

Gly Pro Gly Ile Ser Gly Pro Gly Ser Gly Val Phe Gly Ile Gly Pro

435 440 445

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro

450 455 460

Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala

465 470 475 480

Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly

485 490 495

Pro Gly Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Ser

500 505 510

Ala Ala Ala Ala Ala Gly Ile Tyr Val Phe Gly Pro Gly Val Phe Gly

515 520 525

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly

530 535 540

Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser

545 550 555 560

Gly Val Phe Gly Pro Gly Val Phe Gly Pro Tyr Ala Ser Ala Ala Ala

565 570 575

Ala Ala Gly Pro Gly Ser Gly Val Phe Gly Pro Gly Ala Ser Gly Pro

580 585 590

Gly Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

595 600 605

Gly Ile Asn Gly Pro Gly Ser Gly Val Phe Gly Pro Gly Ile Ser Gly

610 615 620

Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Val Phe Gly Pro Gly Ser

625 630 635 640

Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly Pro Gly Val Phe

645 650 655

Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val

660 665 670

Phe Gly Pro Gly Ala Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro

675 680 685

Gly Val Phe Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Ile Tyr

690 695 700

Gly Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Ser Ala Ala Ala Ala

705 710 715 720

Ala Gly Pro Gly Ser Gly Ile Tyr Gly Ile Gly Pro Tyr Gly Pro Gly

725 730 735

Ala Ser Gly Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Ser Ala

740 745 750

Ser Ala Ala Ala Ala Ala Gly Ser Gly Val Phe Gly Pro Gly Ile Tyr

755 760 765

Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly

770 775 780

Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly Val

785 790 795 800

Phe Gly Pro Gly Val Phe Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala

805 810 815

Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala

820 825 830

Ala Ala Gly Ile Tyr Gly Tyr Gly Pro Gly Val Phe Gly Pro Tyr Gly

835 840 845

Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile Tyr Gly Pro

850 855 860

Gly Val Phe Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Pro Gly

865 870 875 880

Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

885 890 895

Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Tyr Gly Pro Gly Ser

900 905 910

Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala

915 920 925

Ala Ala Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro

930 935 940

Gly Ile Ser Ala Ala Ala Ala Ala Gly Ile Tyr Val Phe Gly Pro Gly

945 950 955 960

Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Val Phe Gly

965 970 975

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile

980 985 990

Tyr Gly Pro Gly Val Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala

995 1000 1005

Gly Ile Tyr Gly Ser Gly Pro Gly Ile Tyr Gly Pro Tyr Gly Pro

1010 1015 1020

Gly Ile Ser Gly Pro Gly Ser Gly Val Phe Gly Ile Gly Pro Tyr

1025 1030 1035

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro

1040 1045 1050

Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala

1055 1060 1065

Ala Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Ala Ser Gly Ile

1070 1075 1080

Asn Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro

1085 1090 1095

Gly Ile Ser Ala Ala Ala Ala Ala Gly Ile Tyr Val Phe Gly Pro

1100 1105 1110

Gly Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala

1115 1120 1125

Ala Gly Ile Tyr Gly Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly

1130 1135 1140

Pro Gly Ile Ser Gly Ser Gly Val Phe Gly Pro Gly Val Phe Gly

1145 1150 1155

Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val

1160 1165 1170

Phe Gly Pro Gly Ala Ser

1175

<210> 33

<211> 601

<212> PRT

<213> Artificial sequence

<220>

<223> PRT888

<400> 33

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Val

1 5 10 15

Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln

20 25 30

Asn Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Gln Ser Gly Gln Tyr

35 40 45

Gly Pro Gly Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Leu Gly

85 90 95

Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly Val

100 105 110

Leu Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser

115 120 125

Gly Pro Gly Val Leu Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Gln Tyr Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Val Leu Gly Pro Gly Gln Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly

195 200 205

Val Leu Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Val Leu Gly

210 215 220

Pro Gly Val Leu Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Val Leu Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Gln Tyr Gly Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro Gly Val

275 280 285

Leu Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Leu

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr

305 310 315 320

Gly Pro Gly Val Leu Gly Pro Gly Gln Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Gln

355 360 365

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Val Leu Gly Pro Gly Val Leu

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Val Leu Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly

405 410 415

Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Gln

420 425 430

Tyr Gly Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser

435 440 445

Gly Pro Gly Ser Gly Val Leu Gly Gln Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro

465 470 475 480

Tyr Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln

500 505 510

Tyr Gly Pro Gly Val Leu Gly Pro Gly Gln Ser Ala Ala Ala Ala Ala

515 520 525

Gly Gln Tyr Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Gln Tyr Gly Ser Gly Pro Gly Val

545 550 555 560

Leu Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Val Leu Gly Pro

565 570 575

Gly Val Leu Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Val Leu Gly Pro Gly Ala Ser

595 600

<210> 34

<211> 601

<212> PRT

<213> Artificial sequence

<220>

<223> PRT965

<400> 34

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Thr

1 5 10 15

Ser Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ala

20 25 30

Asn Gly Pro Gly Ser Gly Thr Ser Gly Pro Gly Ala Ser Gly Ala Tyr

35 40 45

Gly Pro Gly Thr Ser Gly Pro Gly Thr Ser Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Thr Ser Gly

85 90 95

Pro Gly Ala Ser Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro Gly Thr

100 105 110

Ser Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Ala Tyr Gly Ser

115 120 125

Gly Pro Gly Thr Ser Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Ala Tyr Gly Ala Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Thr Ser Gly Pro Gly Ala Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Ala Tyr Gly Ser Gly Pro Gly

195 200 205

Thr Ser Gly Pro Tyr Gly Pro Gly Ala Ser Gly Ser Gly Thr Ser Gly

210 215 220

Pro Gly Thr Ser Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Ala Tyr Gly Tyr Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Ala Asn Gly Pro Gly Ser Gly Ala Tyr Gly Pro Gly Thr

275 280 285

Ser Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Thr Ser

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ala Tyr

305 310 315 320

Gly Pro Gly Thr Ser Gly Pro Gly Ala Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly Ala

355 360 365

Ser Ala Ala Ala Ala Ala Gly Ala Tyr Thr Ser Gly Pro Gly Thr Ser

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Thr Ser Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ala Tyr Gly

405 410 415

Pro Gly Thr Ser Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ala

420 425 430

Tyr Gly Ser Gly Pro Gly Ala Tyr Gly Pro Tyr Gly Pro Gly Ala Ser

435 440 445

Gly Pro Gly Ser Gly Thr Ser Gly Ala Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Ala Tyr Gly Pro Gly Thr Ser Gly Pro

465 470 475 480

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Ala Tyr Gly Pro Gly Ala Ser Gly Ala Asn Gly Pro Gly Ser Gly Ala

500 505 510

Tyr Gly Pro Gly Thr Ser Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

515 520 525

Gly Ala Tyr Thr Ser Gly Pro Gly Thr Ser Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Ala Tyr Gly Ser Gly Pro Gly Thr

545 550 555 560

Ser Gly Pro Tyr Gly Pro Gly Ala Ser Gly Ser Gly Thr Ser Gly Pro

565 570 575

Gly Thr Ser Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Thr Ser Gly Pro Gly Ala Ser

595 600

<210> 35

<211> 601

<212> PRT

<213> Artificial sequence

<220>

<223> PRT889

<400> 35

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Val

1 5 10 15

Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile

20 25 30

Asn Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ile Ser Gly Ile Tyr

35 40 45

Gly Pro Gly Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Leu Gly

85 90 95

Pro Gly Ala Ser Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly Val

100 105 110

Leu Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser

115 120 125

Gly Pro Gly Val Leu Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Ile Tyr Gly Ile Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Val Leu Gly Pro Gly Ile Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly

195 200 205

Val Leu Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly Val Leu Gly

210 215 220

Pro Gly Val Leu Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Val Leu Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Ile Tyr Gly Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Val

275 280 285

Leu Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Leu

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr

305 310 315 320

Gly Pro Gly Val Leu Gly Pro Gly Ile Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ile

355 360 365

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Val Leu Gly Pro Gly Val Leu

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Val Leu Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly

405 410 415

Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ile

420 425 430

Tyr Gly Ser Gly Pro Gly Ile Tyr Gly Pro Tyr Gly Pro Gly Ile Ser

435 440 445

Gly Pro Gly Ser Gly Val Leu Gly Ile Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro

465 470 475 480

Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Ile Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile

500 505 510

Tyr Gly Pro Gly Val Leu Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala

515 520 525

Gly Ile Tyr Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly Val

545 550 555 560

Leu Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly Val Leu Gly Pro

565 570 575

Gly Val Leu Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Val Leu Gly Pro Gly Ala Ser

595 600

<210> 36

<211> 601

<212> PRT

<213> Artificial sequence

<220>

<223> PRT916

<400> 36

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Val

1 5 10 15

Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Leu

20 25 30

Asn Gly Pro Gly Ser Gly Val Ile Gly Pro Gly Leu Ser Gly Leu Tyr

35 40 45

Gly Pro Gly Val Ile Gly Pro Gly Val Ile Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Ile Gly

85 90 95

Pro Gly Ala Ser Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro Gly Val

100 105 110

Ile Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Leu Tyr Gly Ser

115 120 125

Gly Pro Gly Val Ile Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Leu Tyr Gly Leu Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Val Ile Gly Pro Gly Leu Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Leu Tyr Gly Ser Gly Pro Gly

195 200 205

Val Ile Gly Pro Tyr Gly Pro Gly Leu Ser Gly Ser Gly Val Ile Gly

210 215 220

Pro Gly Val Ile Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Val Ile Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Leu Tyr Gly Tyr Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Leu Asn Gly Pro Gly Ser Gly Leu Tyr Gly Pro Gly Val

275 280 285

Ile Gly Pro Gly Leu Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Ile

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Leu Tyr

305 310 315 320

Gly Pro Gly Val Ile Gly Pro Gly Leu Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly Leu

355 360 365

Ser Ala Ala Ala Ala Ala Gly Leu Tyr Val Ile Gly Pro Gly Val Ile

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Val Ile Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Leu Tyr Gly

405 410 415

Pro Gly Val Ile Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Leu

420 425 430

Tyr Gly Ser Gly Pro Gly Leu Tyr Gly Pro Tyr Gly Pro Gly Leu Ser

435 440 445

Gly Pro Gly Ser Gly Val Ile Gly Leu Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Leu Tyr Gly Pro Gly Val Ile Gly Pro

465 470 475 480

Tyr Gly Pro Gly Leu Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Leu Tyr Gly Pro Gly Ala Ser Gly Leu Asn Gly Pro Gly Ser Gly Leu

500 505 510

Tyr Gly Pro Gly Val Ile Gly Pro Gly Leu Ser Ala Ala Ala Ala Ala

515 520 525

Gly Leu Tyr Val Ile Gly Pro Gly Val Ile Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Leu Tyr Gly Ser Gly Pro Gly Val

545 550 555 560

Ile Gly Pro Tyr Gly Pro Gly Leu Ser Gly Ser Gly Val Ile Gly Pro

565 570 575

Gly Val Ile Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Val Ile Gly Pro Gly Ala Ser

595 600

<210> 37

<211> 601

<212> PRT

<213> Artificial sequence

<220>

<223> PRT918

<400> 37

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Val

1 5 10 15

Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile

20 25 30

Asn Gly Pro Gly Ser Gly Val Phe Gly Pro Gly Ile Ser Gly Ile Tyr

35 40 45

Gly Pro Gly Val Phe Gly Pro Gly Val Phe Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Phe Gly

85 90 95

Pro Gly Ala Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Val

100 105 110

Phe Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser

115 120 125

Gly Pro Gly Val Phe Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Ile Tyr Gly Ile Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Val Phe Gly Pro Gly Ile Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly

195 200 205

Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly Val Phe Gly

210 215 220

Pro Gly Val Phe Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Ile Tyr Gly Tyr Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Val

275 280 285

Phe Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Phe

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr

305 310 315 320

Gly Pro Gly Val Phe Gly Pro Gly Ile Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile

355 360 365

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Val Phe Gly Pro Gly Val Phe

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Val Phe Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly

405 410 415

Pro Gly Val Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ile

420 425 430

Tyr Gly Ser Gly Pro Gly Ile Tyr Gly Pro Tyr Gly Pro Gly Ile Ser

435 440 445

Gly Pro Gly Ser Gly Val Phe Gly Ile Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro

465 470 475 480

Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Ile Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile

500 505 510

Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala

515 520 525

Gly Ile Tyr Val Phe Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly Val

545 550 555 560

Phe Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly Val Phe Gly Pro

565 570 575

Gly Val Phe Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Val Phe Gly Pro Gly Ala Ser

595 600

<210> 38

<211> 576

<212> PRT

<213> Artificial sequence

<220>

<223> PRT699

<400> 38

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Val

1 5 10 15

Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala

20 25 30

Gly Ser Asn Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Gln Ser Gly

35 40 45

Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser

50 55 60

Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly

65 70 75 80

Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro

85 90 95

Gly Ser Gly Val Leu Gly Pro Gly Ala Ser Gly Gln Tyr Gly Pro Gly

100 105 110

Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser Ser Ala Ala Ala Ala

115 120 125

Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly Val Leu Gly Pro Tyr

130 135 140

Gly Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Gln Tyr

145 150 155 160

Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Gln Tyr Gly

165 170 175

Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala

180 185 190

Gly Ser Gly Val Leu Gly Pro Gly Gln Tyr Gly Pro Tyr Ala Ser Ala

195 200 205

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly Val Leu

210 215 220

Gly Pro Tyr Gly Pro Gly Gln Ser Gly Ser Gly Val Leu Gly Pro Gly

225 230 235 240

Val Leu Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro

245 250 255

Gly Val Leu Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

260 265 270

Ala Ala Gly Ser Tyr Gly Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly

275 280 285

Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser Gly Gln Tyr Gly Pro

290 295 300

Gly Val Leu Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly

305 310 315 320

Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala

325 330 335

Ala Ala Ala Gly Ser Tyr Gly Pro Gly Val Leu Gly Pro Gly Gln Tyr

340 345 350

Gly Pro Gly Ser Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

355 360 365

Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Val

370 375 380

Leu Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala

385 390 395 400

Gly Ser Tyr Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

405 410 415

Ala Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala

420 425 430

Ala Ala Ala Ala Ala Ala Gly Pro Gly Gln Tyr Gly Pro Gly Val Leu

435 440 445

Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly

450 455 460

Ser Gly Pro Gly Gln Tyr Gly Pro Tyr Gly Pro Gly Gln Ser Gly Pro

465 470 475 480

Gly Ser Gly Val Leu Gly Gln Gly Pro Tyr Gly Pro Gly Ala Ser Ala

485 490 495

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Val Leu Gly Pro

500 505 510

Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

515 520 525

Ser Gly Gln Tyr Gly Pro Gly Ala Ser Gly Gln Asn Gly Pro Gly Ser

530 535 540

Gly Gln Tyr Gly Pro Gly Val Leu Gly Pro Gly Pro Ser Ala Ala Ala

545 550 555 560

Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ala Ser

565 570 575

<210> 39

<211> 576

<212> PRT

<213> Artificial sequence

<220>

<223> PRT698

<400> 39

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Val

1 5 10 15

Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Ala Ala

20 25 30

Gly Ser Asn Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ile Ser Gly

35 40 45

Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser

50 55 60

Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly Pro Gly

65 70 75 80

Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro

85 90 95

Gly Ser Gly Val Leu Gly Pro Gly Ala Ser Gly Ile Tyr Gly Pro Gly

100 105 110

Val Leu Gly Pro Gly Val Leu Gly Pro Gly Ser Ser Ala Ala Ala Ala

115 120 125

Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly Val Leu Gly Pro Tyr

130 135 140

Gly Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Ile Tyr

145 150 155 160

Gly Ile Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Ile Tyr Gly

165 170 175

Pro Gly Val Leu Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala

180 185 190

Gly Ser Gly Val Leu Gly Pro Gly Ile Tyr Gly Pro Tyr Ala Ser Ala

195 200 205

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Ser Gly Pro Gly Val Leu

210 215 220

Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly Val Leu Gly Pro Gly

225 230 235 240

Val Leu Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro

245 250 255

Gly Val Leu Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

260 265 270

Ala Ala Gly Ser Tyr Gly Tyr Gly Pro Gly Val Leu Gly Pro Tyr Gly

275 280 285

Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile Tyr Gly Pro

290 295 300

Gly Val Leu Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly

305 310 315 320

Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala

325 330 335

Ala Ala Ala Gly Ser Tyr Gly Pro Gly Val Leu Gly Pro Gly Ile Tyr

340 345 350

Gly Pro Gly Ser Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

355 360 365

Ser Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Val

370 375 380

Leu Gly Pro Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala

385 390 395 400

Gly Ser Tyr Val Leu Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly

405 410 415

Ala Ser Gly Pro Gly Val Leu Gly Pro Tyr Gly Pro Gly Ala Ser Ala

420 425 430

Ala Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly Pro Gly Val Leu

435 440 445

Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly

450 455 460

Ser Gly Pro Gly Ile Tyr Gly Pro Tyr Gly Pro Gly Ile Ser Gly Pro

465 470 475 480

Gly Ser Gly Val Leu Gly Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala

485 490 495

Ala Ala Ala Ala Ala Ala Gly Ser Tyr Gly Pro Gly Val Leu Gly Pro

500 505 510

Tyr Gly Pro Gly Pro Ser Ala Ala Ala Ala Ala Ala Ala Gly Pro Gly

515 520 525

Ser Gly Ile Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly Ser

530 535 540

Gly Ile Tyr Gly Pro Gly Val Leu Gly Pro Gly Pro Ser Ala Ala Ala

545 550 555 560

Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Leu Gly Pro Gly Ala Ser

565 570 575

<210> 40

<211> 1190

<212> PRT

<213> Artificial sequence

<220>

<223> PRT966

<400> 40

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Val

1 5 10 15

Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile

20 25 30

Asn Gly Pro Gly Ser Gly Val Phe Gly Pro Gly Ile Ser Gly Ile Tyr

35 40 45

Gly Pro Gly Val Phe Gly Pro Gly Val Phe Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Phe Gly

85 90 95

Pro Gly Ala Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Val

100 105 110

Phe Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser

115 120 125

Gly Pro Gly Val Phe Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Ile Tyr Gly Ile Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Val Phe Gly Pro Gly Ile Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly

195 200 205

Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly Val Phe Gly

210 215 220

Pro Gly Val Phe Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Ile Tyr Gly Tyr Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Val

275 280 285

Phe Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Phe

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr

305 310 315 320

Gly Pro Gly Val Phe Gly Pro Gly Ile Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile

355 360 365

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Val Phe Gly Pro Gly Val Phe

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Val Phe Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly

405 410 415

Pro Gly Val Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ile

420 425 430

Tyr Gly Ser Gly Pro Gly Ile Tyr Gly Pro Tyr Gly Pro Gly Ile Ser

435 440 445

Gly Pro Gly Ser Gly Val Phe Gly Ile Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro

465 470 475 480

Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Ile Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly Ser Gly Ile

500 505 510

Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala

515 520 525

Gly Ile Tyr Val Phe Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly Val

545 550 555 560

Phe Gly Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly Val Phe Gly Pro

565 570 575

Gly Val Phe Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Val Phe Gly Pro Gly Ala Ser Gly Pro Gly Val Phe Gly Pro

595 600 605

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Asn Gly Pro

610 615 620

Gly Ser Gly Val Phe Gly Pro Gly Ile Ser Gly Ile Tyr Gly Pro Gly

625 630 635 640

Val Phe Gly Pro Gly Val Phe Gly Pro Gly Ser Ser Ala Ala Ala Ala

645 650 655

Ala Gly Pro Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Ser Ala Ser

660 665 670

Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Phe Gly Pro Gly Ala

675 680 685

Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Val Phe Gly Pro

690 695 700

Gly Ser Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly

705 710 715 720

Val Phe Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser

725 730 735

Gly Ile Tyr Gly Ile Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly

740 745 750

Ile Tyr Gly Pro Gly Val Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala

755 760 765

Ala Gly Ser Gly Val Phe Gly Pro Gly Ile Tyr Gly Pro Tyr Ala Ser

770 775 780

Ala Ala Ala Ala Ala Gly Ile Tyr Gly Ser Gly Pro Gly Val Phe Gly

785 790 795 800

Pro Tyr Gly Pro Gly Ile Ser Gly Ser Gly Val Phe Gly Pro Gly Val

805 810 815

Phe Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Phe

820 825 830

Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Ile Tyr

835 840 845

Gly Tyr Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ala Ser Gly

850 855 860

Ile Asn Gly Pro Gly Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro

865 870 875 880

Gly Ile Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Phe Gly Pro Tyr

885 890 895

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro Gly

900 905 910

Val Phe Gly Pro Gly Ile Tyr Gly Pro Gly Ser Ser Gly Pro Gly Val

915 920 925

Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Ile

930 935 940

Tyr Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser Ala Ala

945 950 955 960

Ala Ala Ala Gly Ile Tyr Val Phe Gly Pro Gly Val Phe Gly Pro Tyr

965 970 975

Gly Pro Gly Ala Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly

980 985 990

Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile Tyr Gly Pro Gly Val

995 1000 1005

Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly

1010 1015 1020

Ser Gly Pro Gly Ile Tyr Gly Pro Tyr Gly Pro Gly Ile Ser Gly

1025 1030 1035

Pro Gly Ser Gly Val Phe Gly Ile Gly Pro Tyr Gly Pro Gly Ala

1040 1045 1050

Ser Ala Ala Ala Ala Ala Gly Ile Tyr Gly Pro Gly Val Phe Gly

1055 1060 1065

Pro Tyr Gly Pro Gly Ile Ser Ala Ala Ala Ala Ala Gly Pro Gly

1070 1075 1080

Ser Gly Ile Tyr Gly Pro Gly Ala Ser Gly Ile Asn Gly Pro Gly

1085 1090 1095

Ser Gly Ile Tyr Gly Pro Gly Val Phe Gly Pro Gly Ile Ser Ala

1100 1105 1110

Ala Ala Ala Ala Gly Ile Tyr Val Phe Gly Pro Gly Val Phe Gly

1115 1120 1125

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Ile Tyr

1130 1135 1140

Gly Ser Gly Pro Gly Val Phe Gly Pro Tyr Gly Pro Gly Ile Ser

1145 1150 1155

Gly Ser Gly Val Phe Gly Pro Gly Val Phe Gly Pro Tyr Ala Ser

1160 1165 1170

Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Phe Gly Pro Gly

1175 1180 1185

Ala Ser

1190

<210> 41

<211> 590

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT917

<400> 41

Met Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Val Asn Gly Pro Gly Ser Gly Leu Ile Gly Pro Gly

20 25 30

Val Ser Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro Gly Leu Ile Gly

35 40 45

Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Tyr Gly Pro

50 55 60

Gly Leu Ile Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

65 70 75 80

Ser Gly Leu Ile Gly Pro Gly Ala Ser Gly Val Tyr Gly Pro Gly Leu

85 90 95

Ile Gly Pro Gly Leu Ile Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

100 105 110

Gly Val Tyr Gly Ser Gly Pro Gly Leu Ile Gly Pro Tyr Gly Ser Ala

115 120 125

Ala Ala Ala Ala Gly Pro Gly Ser Gly Val Tyr Gly Val Gly Pro Tyr

130 135 140

Gly Pro Gly Ala Ser Gly Pro Gly Val Tyr Gly Pro Gly Leu Ile Gly

145 150 155 160

Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Leu Ile Gly Pro

165 170 175

Gly Val Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Val Tyr

180 185 190

Gly Ser Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly Val Ser Gly

195 200 205

Ser Gly Leu Ile Gly Pro Gly Leu Ile Gly Pro Tyr Ala Ser Ala Ala

210 215 220

Ala Ala Ala Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly Ser Ser

225 230 235 240

Ala Ala Ala Ala Ala Gly Val Tyr Gly Tyr Gly Pro Gly Leu Ile Gly

245 250 255

Pro Tyr Gly Pro Gly Ala Ser Gly Val Asn Gly Pro Gly Ser Gly Val

260 265 270

Tyr Gly Pro Gly Leu Ile Gly Pro Gly Val Ser Ala Ala Ala Ala Ala

275 280 285

Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala

290 295 300

Ala Ala Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro Gly Val Tyr Gly

305 310 315 320

Pro Gly Ser Ser Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly Ser

325 330 335

Ser Ala Ala Ala Ala Ala Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro

340 345 350

Tyr Gly Pro Gly Val Ser Ala Ala Ala Ala Ala Gly Val Tyr Leu Ile

355 360 365

Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly

370 375 380

Leu Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly

385 390 395 400

Pro Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro Ser Ala Ser Ala Ala

405 410 415

Ala Ala Ala Gly Val Tyr Gly Ser Gly Pro Gly Val Tyr Gly Pro Tyr

420 425 430

Gly Pro Gly Val Ser Gly Pro Gly Ser Gly Leu Ile Gly Val Gly Pro

435 440 445

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Val Tyr Gly Pro

450 455 460

Gly Leu Ile Gly Pro Tyr Gly Pro Gly Val Ser Ala Ala Ala Ala Ala

465 470 475 480

Gly Pro Gly Ser Gly Val Tyr Gly Pro Gly Ala Ser Gly Val Asn Gly

485 490 495

Pro Gly Ser Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro Gly Val Ser

500 505 510

Ala Ala Ala Ala Ala Gly Val Tyr Leu Ile Gly Pro Gly Leu Ile Gly

515 520 525

Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Val Tyr Gly

530 535 540

Ser Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly Val Ser Gly Ser

545 550 555 560

Gly Leu Ile Gly Pro Gly Leu Ile Gly Pro Tyr Ala Ser Ala Ala Ala

565 570 575

Ala Ala Gly Pro Gly Ser Gly Leu Ile Gly Pro Gly Ala Ser

580 585 590

<210> 42

<211> 587

<212> PRT

<213> Artificial sequence

<220>

<223> Met-PRT1028

<400> 42

Met Gly Pro Gly Ile Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala

1 5 10 15

Ala Ala Ala Gly Thr Gly Pro Gly Ser Gly Ile Phe Gly Pro Gly Thr

20 25 30

Ser Gly Thr Tyr Gly Pro Gly Ile Phe Gly Pro Gly Ile Phe Gly Pro

35 40 45

Gly Ser Ser Ala Ala Ala Ala Ala Gly Pro Gly Thr Tyr Gly Pro Gly

50 55 60

Ile Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser

65 70 75 80

Gly Ile Phe Gly Pro Gly Ala Ser Gly Thr Tyr Gly Pro Gly Ile Phe

85 90 95

Gly Pro Gly Ile Phe Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly

100 105 110

Thr Tyr Gly Ser Gly Pro Gly Ile Phe Gly Pro Tyr Gly Ser Ala Ala

115 120 125

Ala Ala Ala Gly Pro Gly Ser Gly Thr Tyr Gly Thr Gly Pro Tyr Gly

130 135 140

Pro Gly Ala Ser Gly Pro Gly Thr Tyr Gly Pro Gly Ile Phe Gly Pro

145 150 155 160

Ser Ala Ser Ala Ala Ala Ala Ala Gly Ser Gly Ile Phe Gly Pro Gly

165 170 175

Thr Tyr Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Thr Tyr Gly

180 185 190

Ser Gly Pro Gly Ile Phe Gly Pro Tyr Gly Pro Gly Thr Ser Gly Ser

195 200 205

Gly Ile Phe Gly Pro Gly Ile Phe Gly Pro Tyr Ala Ser Ala Ala Ala

210 215 220

Ala Ala Gly Pro Gly Ile Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala

225 230 235 240

Ala Ala Ala Ala Gly Thr Tyr Gly Tyr Gly Pro Gly Ile Phe Gly Pro

245 250 255

Tyr Gly Pro Gly Ala Ser Gly Thr Gly Pro Gly Ser Gly Thr Tyr Gly

260 265 270

Pro Gly Ile Phe Gly Pro Gly Thr Ser Ala Ala Ala Ala Ala Gly Pro

275 280 285

Gly Ile Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala

290 295 300

Gly Thr Tyr Gly Pro Gly Ile Phe Gly Pro Gly Thr Tyr Gly Pro Gly

305 310 315 320

Ser Ser Gly Pro Gly Ile Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala

325 330 335

Ala Ala Ala Ala Gly Thr Tyr Gly Pro Gly Ile Phe Gly Pro Tyr Gly

340 345 350

Pro Gly Thr Ser Ala Ala Ala Ala Ala Gly Thr Tyr Ile Phe Gly Pro

355 360 365

Gly Ile Phe Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Ile Phe

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

385 390 395 400

Thr Tyr Gly Pro Gly Ile Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala

405 410 415

Ala Gly Thr Tyr Gly Ser Gly Pro Gly Thr Tyr Gly Pro Tyr Gly Pro

420 425 430

Gly Thr Ser Gly Pro Gly Ser Gly Ile Phe Gly Thr Gly Pro Tyr Gly

435 440 445

Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Thr Tyr Gly Pro Gly Ile

450 455 460

Phe Gly Pro Tyr Gly Pro Gly Thr Ser Ala Ala Ala Ala Ala Gly Pro

465 470 475 480

Gly Ser Gly Thr Tyr Gly Pro Gly Ala Ser Gly Thr Gly Pro Gly Ser

485 490 495

Gly Thr Tyr Gly Pro Gly Ile Phe Gly Pro Gly Thr Ser Ala Ala Ala

500 505 510

Ala Ala Gly Thr Tyr Ile Phe Gly Pro Gly Ile Phe Gly Pro Tyr Gly

515 520 525

Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Thr Tyr Gly Ser Gly Pro

530 535 540

Gly Ile Phe Gly Pro Tyr Gly Pro Gly Thr Ser Gly Ser Gly Ile Phe

545 550 555 560

Gly Pro Gly Ile Phe Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly

565 570 575

Pro Gly Ser Gly Ile Phe Gly Pro Gly Ala Ser

580 585

<210> 43

<211> 601

<212> PRT

<213> Artificial sequence

<220>

<223> PRT917

<400> 43

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Leu

1 5 10 15

Ile Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Val

20 25 30

Asn Gly Pro Gly Ser Gly Leu Ile Gly Pro Gly Val Ser Gly Val Tyr

35 40 45

Gly Pro Gly Leu Ile Gly Pro Gly Leu Ile Gly Pro Gly Ser Ser Ala

50 55 60

Ala Ala Ala Ala Gly Pro Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro

65 70 75 80

Ser Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Leu Ile Gly

85 90 95

Pro Gly Ala Ser Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro Gly Leu

100 105 110

Ile Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Val Tyr Gly Ser

115 120 125

Gly Pro Gly Leu Ile Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly

130 135 140

Pro Gly Ser Gly Val Tyr Gly Val Gly Pro Tyr Gly Pro Gly Ala Ser

145 150 155 160

Gly Pro Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro Ser Ala Ser Ala

165 170 175

Ala Ala Ala Ala Gly Ser Gly Leu Ile Gly Pro Gly Val Tyr Gly Pro

180 185 190

Tyr Ala Ser Ala Ala Ala Ala Ala Gly Val Tyr Gly Ser Gly Pro Gly

195 200 205

Leu Ile Gly Pro Tyr Gly Pro Gly Val Ser Gly Ser Gly Leu Ile Gly

210 215 220

Pro Gly Leu Ile Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro

225 230 235 240

Gly Leu Ile Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala

245 250 255

Gly Val Tyr Gly Tyr Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly

260 265 270

Ala Ser Gly Val Asn Gly Pro Gly Ser Gly Val Tyr Gly Pro Gly Leu

275 280 285

Ile Gly Pro Gly Val Ser Ala Ala Ala Ala Ala Gly Pro Gly Leu Ile

290 295 300

Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Val Tyr

305 310 315 320

Gly Pro Gly Leu Ile Gly Pro Gly Val Tyr Gly Pro Gly Ser Ser Gly

325 330 335

Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala

340 345 350

Ala Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly Val

355 360 365

Ser Ala Ala Ala Ala Ala Gly Val Tyr Leu Ile Gly Pro Gly Leu Ile

370 375 380

Gly Pro Tyr Gly Pro Gly Ala Ser Gly Pro Gly Leu Ile Gly Pro Tyr

385 390 395 400

Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Val Tyr Gly

405 410 415

Pro Gly Leu Ile Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Val

420 425 430

Tyr Gly Ser Gly Pro Gly Val Tyr Gly Pro Tyr Gly Pro Gly Val Ser

435 440 445

Gly Pro Gly Ser Gly Leu Ile Gly Val Gly Pro Tyr Gly Pro Gly Ala

450 455 460

Ser Ala Ala Ala Ala Ala Gly Val Tyr Gly Pro Gly Leu Ile Gly Pro

465 470 475 480

Tyr Gly Pro Gly Val Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly

485 490 495

Val Tyr Gly Pro Gly Ala Ser Gly Val Asn Gly Pro Gly Ser Gly Val

500 505 510

Tyr Gly Pro Gly Leu Ile Gly Pro Gly Val Ser Ala Ala Ala Ala Ala

515 520 525

Gly Val Tyr Leu Ile Gly Pro Gly Leu Ile Gly Pro Tyr Gly Pro Gly

530 535 540

Ala Ser Ala Ala Ala Ala Ala Gly Val Tyr Gly Ser Gly Pro Gly Leu

545 550 555 560

Ile Gly Pro Tyr Gly Pro Gly Val Ser Gly Ser Gly Leu Ile Gly Pro

565 570 575

Gly Leu Ile Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

580 585 590

Ser Gly Leu Ile Gly Pro Gly Ala Ser

595 600

<210> 44

<211> 598

<212> PRT

<213> Artificial sequence

<220>

<223> PRT1028

<400> 44

Met His His His His His His Ser Ser Gly Ser Ser Gly Pro Gly Ile

1 5 10 15

Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Thr

20 25 30

Gly Pro Gly Ser Gly Ile Phe Gly Pro Gly Thr Ser Gly Thr Tyr Gly

35 40 45

Pro Gly Ile Phe Gly Pro Gly Ile Phe Gly Pro Gly Ser Ser Ala Ala

50 55 60

Ala Ala Ala Gly Pro Gly Thr Tyr Gly Pro Gly Ile Phe Gly Pro Ser

65 70 75 80

Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Ile Phe Gly Pro

85 90 95

Gly Ala Ser Gly Thr Tyr Gly Pro Gly Ile Phe Gly Pro Gly Ile Phe

100 105 110

Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly Thr Tyr Gly Ser Gly

115 120 125

Pro Gly Ile Phe Gly Pro Tyr Gly Ser Ala Ala Ala Ala Ala Gly Pro

130 135 140

Gly Ser Gly Thr Tyr Gly Thr Gly Pro Tyr Gly Pro Gly Ala Ser Gly

145 150 155 160

Pro Gly Thr Tyr Gly Pro Gly Ile Phe Gly Pro Ser Ala Ser Ala Ala

165 170 175

Ala Ala Ala Gly Ser Gly Ile Phe Gly Pro Gly Thr Tyr Gly Pro Tyr

180 185 190

Ala Ser Ala Ala Ala Ala Ala Gly Thr Tyr Gly Ser Gly Pro Gly Ile

195 200 205

Phe Gly Pro Tyr Gly Pro Gly Thr Ser Gly Ser Gly Ile Phe Gly Pro

210 215 220

Gly Ile Phe Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly

225 230 235 240

Ile Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly

245 250 255

Thr Tyr Gly Tyr Gly Pro Gly Ile Phe Gly Pro Tyr Gly Pro Gly Ala

260 265 270

Ser Gly Thr Gly Pro Gly Ser Gly Thr Tyr Gly Pro Gly Ile Phe Gly

275 280 285

Pro Gly Thr Ser Ala Ala Ala Ala Ala Gly Pro Gly Ile Phe Gly Pro

290 295 300

Tyr Gly Pro Gly Ala Ser Ala Ala Ala Ala Ala Gly Thr Tyr Gly Pro

305 310 315 320

Gly Ile Phe Gly Pro Gly Thr Tyr Gly Pro Gly Ser Ser Gly Pro Gly

325 330 335

Ile Phe Gly Pro Tyr Gly Pro Gly Ser Ser Ala Ala Ala Ala Ala Gly

340 345 350

Thr Tyr Gly Pro Gly Ile Phe Gly Pro Tyr Gly Pro Gly Thr Ser Ala

355 360 365

Ala Ala Ala Ala Gly Thr Tyr Ile Phe Gly Pro Gly Ile Phe Gly Pro

370 375 380

Tyr Gly Pro Gly Ala Ser Gly Pro Gly Ile Phe Gly Pro Tyr Gly Pro

385 390 395 400

Gly Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Thr Tyr Gly Pro Gly

405 410 415

Ile Phe Gly Pro Ser Ala Ser Ala Ala Ala Ala Ala Gly Thr Tyr Gly

420 425 430

Ser Gly Pro Gly Thr Tyr Gly Pro Tyr Gly Pro Gly Thr Ser Gly Pro

435 440 445

Gly Ser Gly Ile Phe Gly Thr Gly Pro Tyr Gly Pro Gly Ala Ser Ala

450 455 460

Ala Ala Ala Ala Gly Thr Tyr Gly Pro Gly Ile Phe Gly Pro Tyr Gly

465 470 475 480

Pro Gly Thr Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Thr Tyr

485 490 495

Gly Pro Gly Ala Ser Gly Thr Gly Pro Gly Ser Gly Thr Tyr Gly Pro

500 505 510

Gly Ile Phe Gly Pro Gly Thr Ser Ala Ala Ala Ala Ala Gly Thr Tyr

515 520 525

Ile Phe Gly Pro Gly Ile Phe Gly Pro Tyr Gly Pro Gly Ala Ser Ala

530 535 540

Ala Ala Ala Ala Gly Thr Tyr Gly Ser Gly Pro Gly Ile Phe Gly Pro

545 550 555 560

Tyr Gly Pro Gly Thr Ser Gly Ser Gly Ile Phe Gly Pro Gly Ile Phe

565 570 575

Gly Pro Tyr Ala Ser Ala Ala Ala Ala Ala Gly Pro Gly Ser Gly Ile

580 585 590

Phe Gly Pro Gly Ala Ser

595

<210> 45

<211> 252

<212> PRT

<213> Artificial sequence

<220>

<223> Collagen-4 type-Kai

<400> 45

Met His His His His His His Ser Ser Gly Ser Ser Lys Asp Gly Val

1 5 10 15

Pro Gly Phe Pro Gly Ser Glu Gly Val Lys Gly Asn Arg Gly Phe Pro

20 25 30

Gly Leu Met Gly Glu Asp Gly Ile Lys Gly Gln Lys Gly Asp Ile Gly

35 40 45

Pro Pro Gly Phe Arg Gly Pro Thr Glu Tyr Tyr Asp Thr Tyr Gln Glu

50 55 60

Lys Gly Asp Glu Gly Thr Pro Gly Pro Pro Gly Pro Arg Gly Ala Arg

65 70 75 80

Gly Pro Gln Gly Pro Ser Gly Pro Pro Gly Val Pro Gly Ser Pro Gly

85 90 95

Ser Ser Arg Pro Gly Leu Arg Gly Ala Pro Gly Trp Pro Gly Leu Lys

100 105 110

Gly Ser Lys Gly Glu Arg Gly Arg Pro Gly Lys Asp Ala Met Gly Thr

115 120 125

Pro Gly Ser Pro Gly Cys Ala Gly Ser Pro Gly Leu Pro Gly Ser Pro

130 135 140

Gly Pro Pro Gly Pro Pro Gly Asp Ile Val Phe Arg Lys Gly Pro Pro

145 150 155 160

Gly Asp His Gly Leu Pro Gly Tyr Leu Gly Ser Pro Gly Ile Pro Gly

165 170 175

Val Asp Gly Pro Lys Gly Glu Pro Gly Leu Leu Cys Thr Gln Cys Pro

180 185 190

Tyr Ile Pro Gly Pro Pro Gly Leu Pro Gly Leu Pro Gly Leu His Gly

195 200 205

Val Lys Gly Ile Pro Gly Arg Gln Gly Ala Ala Gly Leu Lys Gly Ser

210 215 220

Pro Gly Ser Pro Gly Asn Thr Gly Leu Pro Gly Phe Pro Gly Phe Pro

225 230 235 240

Gly Ala Gln Gly Asp Pro Gly Leu Lys Gly Glu Lys

245 250

<210> 46

<211> 310

<212> PRT

<213> Artificial sequence

<220>

<223> Arthropoda elastin-Kai

<400> 46

Met His His His His His His Pro Glu Pro Pro Val Asn Ser Tyr Leu

1 5 10 15

Pro Pro Ser Asp Ser Tyr Gly Ala Pro Gly Gln Ser Gly Pro Gly Gly

20 25 30

Arg Pro Ser Asp Ser Tyr Gly Ala Pro Gly Gly Gly Asn Gly Gly Arg

35 40 45

Pro Ser Asp Ser Tyr Gly Ala Pro Gly Gln Gly Gln Gly Gln Gly Gln

50 55 60

Gly Gln Gly Gly Tyr Ala Gly Lys Pro Ser Asp Ser Tyr Gly Ala Pro

65 70 75 80

Gly Gly Gly Asp Gly Asn Gly Gly Arg Pro Ser Ser Ser Tyr Gly Ala

85 90 95

Pro Gly Gly Gly Asn Gly Gly Arg Pro Ser Asp Thr Tyr Gly Ala Pro

100 105 110

Gly Gly Gly Asn Gly Gly Arg Pro Ser Asp Thr Tyr Gly Ala Pro Gly

115 120 125

Gly Gly Gly Asn Gly Asn Gly Gly Arg Pro Ser Ser Ser Tyr Gly Ala

130 135 140

Pro Gly Gln Gly Gln Gly Asn Gly Asn Gly Gly Arg Pro Ser Ser Ser

145 150 155 160

Tyr Gly Ala Pro Gly Gly Gly Asn Gly Gly Arg Pro Ser Asp Thr Tyr

165 170 175

Gly Ala Pro Gly Gly Gly Asn Gly Gly Arg Pro Ser Asp Thr Tyr Gly

180 185 190

Ala Pro Gly Gly Gly Asn Asn Gly Gly Arg Pro Ser Ser Ser Tyr Gly

195 200 205

Ala Pro Gly Gly Gly Asn Gly Gly Arg Pro Ser Asp Thr Tyr Gly Ala

210 215 220

Pro Gly Gly Gly Asn Gly Asn Gly Ser Gly Gly Arg Pro Ser Ser Ser

225 230 235 240

Tyr Gly Ala Pro Gly Gln Gly Gln Gly Gly Phe Gly Gly Arg Pro Ser

245 250 255

Asp Ser Tyr Gly Ala Pro Gly Gln Asn Gln Lys Pro Ser Asp Ser Tyr

260 265 270

Gly Ala Pro Gly Ser Gly Asn Gly Asn Gly Gly Arg Pro Ser Ser Ser

275 280 285

Tyr Gly Ala Pro Gly Ser Gly Pro Gly Gly Arg Pro Ser Asp Ser Tyr

290 295 300

Gly Pro Pro Ala Ser Gly

305 310

<210> 47

<211> 282

<212> PRT

<213> Artificial sequence

<220>

<223> Elastin deficiency

<400> 47

Met His His His His His His Ser Ser Gly Ser Ser Leu Gly Val Ser

1 5 10 15

Ala Gly Ala Val Val Pro Gln Pro Gly Ala Gly Val Lys Pro Gly Lys

20 25 30

Val Pro Gly Val Gly Leu Pro Gly Val Tyr Pro Gly Gly Val Leu Pro

35 40 45

Gly Ala Arg Phe Pro Gly Val Gly Val Leu Pro Gly Val Pro Thr Gly

50 55 60

Ala Gly Val Lys Pro Lys Ala Pro Gly Val Gly Gly Ala Phe Ala Gly

65 70 75 80

Ile Pro Gly Val Gly Pro Phe Gly Gly Pro Gln Pro Gly Val Pro Leu

85 90 95

Gly Tyr Pro Ile Lys Ala Pro Lys Leu Pro Gly Gly Tyr Gly Leu Pro

100 105 110

Tyr Thr Thr Gly Lys Leu Pro Tyr Gly Tyr Gly Pro Gly Gly Val Ala

115 120 125

Gly Ala Ala Gly Lys Ala Gly Tyr Pro Thr Gly Thr Gly Val Gly Pro

130 135 140

Gln Ala Ala Ala Ala Ala Ala Ala Lys Ala Ala Ala Lys Phe Gly Ala

145 150 155 160

Gly Ala Ala Gly Val Leu Pro Gly Val Gly Gly Ala Gly Val Pro Gly

165 170 175

Val Pro Gly Ala Ile Pro Gly Ile Gly Gly Ile Ala Gly Val Gly Thr

180 185 190

Pro Ala Ala Ala Ala Ala Ala Ala Ala Ala Ala Lys Ala Ala Lys Tyr

195 200 205

Gly Ala Ala Ala Gly Leu Val Pro Gly Gly Pro Gly Phe Gly Pro Gly

210 215 220

Val Val Gly Val Pro Gly Ala Gly Val Pro Gly Val Gly Val Pro Gly

225 230 235 240

Ala Gly Ile Pro Val Val Pro Gly Ala Gly Ile Pro Gly Ala Ala Val

245 250 255

Pro Gly Val Val Ser Pro Glu Ala Ala Ala Lys Ala Ala Ala Lys Ala

260 265 270

Ala Lys Tyr Gly Ala Arg Pro Gly Val Gly

275 280

<210> 48

<211> 468

<212> PRT

<213> Artificial sequence

<220>

<223> Type I Keratin 26

<400> 48

Met Ser Phe Arg Leu Ser Gly Val Ser Arg Arg Leu Cys Ser Gln Ala

1 5 10 15

Gly Thr Gly Arg Leu Thr Gly Gly Arg Thr Gly Phe Arg Ala Gly Asn

20 25 30

Val Cys Ser Gly Leu Gly Ala Gly Ser Ser Phe Ser Gly Pro Leu Gly

35 40 45

Ser Val Ser Ser Lys Gly Ser Phe Ser His Gly Gly Gly Gly Leu Gly

50 55 60

Ser Gly Val Cys Thr Gly Phe Leu Glu Asn Glu His Gly Leu Leu Pro

65 70 75 80

Gly Asn Glu Lys Val Thr Leu Gln Asn Leu Asn Asp Arg Leu Ala Ser

85 90 95

Tyr Leu Asp His Val Cys Thr Leu Glu Glu Ala Asn Ala Asp Leu Glu

100 105 110

Gln Lys Ile Lys Gly Trp Tyr Glu Lys Tyr Gly Pro Gly Ser Gly Arg

115 120 125

Gln Leu Ala His Asp Tyr Ser Lys Tyr Phe Ser Val Thr Glu Asp Leu

130 135 140

Lys Arg Gln Ile Ile Ser Val Thr Thr Cys Asn Ala Ser Ile Val Leu

145 150 155 160

Gln Asn Glu Asn Ala Arg Leu Thr Ala Asp Asp Phe Arg Leu Lys Cys

165 170 175

Glu Asn Glu Leu Ala Leu His Gln Ser Val Glu Ala Asp Ile Asn Gly

180 185 190

Leu His Arg Val Met Asp Glu Leu Thr Leu Cys Thr Ser Asp Leu Glu

195 200 205

Met Gln Cys Glu Ala Leu Ser Glu Glu Leu Thr Tyr Leu Lys Lys Asn

210 215 220

His Gln Glu Glu Met Lys Val Met Gln Gly Ala Ala Arg Gly Asn Val

225 230 235 240

Asn Val Glu Ile Asn Ala Ala Pro Gly Val Asp Leu Thr Val Leu Leu

245 250 255

Asn Asn Met Arg Ala Glu Tyr Glu Asp Leu Ala Glu Gln Asn His Glu

260 265 270

Asp Ala Glu Ala Trp Phe Ser Glu Lys Ser Thr Ser Leu His Gln Gln

275 280 285

Ile Ser Asp Asp Ala Gly Ala Ala Met Ala Ala Arg Asn Glu Leu Met

290 295 300

Glu Leu Lys Arg Asn Leu Gln Thr Leu Glu Ile Glu Leu Gln Ser Leu

305 310 315 320

Leu Ala Met Lys His Ser Tyr Glu Cys Ser Leu Ala Glu Thr Glu Ser

325 330 335

Asn Tyr Cys His Gln Leu Gln Gln Ile Gln Glu Gln Ile Gly Ala Met

340 345 350

Glu Asp Gln Leu Gln Gln Ile Arg Met Glu Thr Glu Gly Gln Lys Leu

355 360 365

Glu His Glu Arg Leu Leu Asp Val Lys Ile Phe Leu Glu Lys Glu Ile

370 375 380

Glu Met Tyr Cys Lys Leu Ile Asp Gly Glu Gly Arg Lys Ser Lys Ser

385 390 395 400

Thr Cys Tyr Lys Ser Glu Gly Arg Gly Pro Lys Asn Ser Glu Asn Gln

405 410 415

Val Lys Asp Ser Lys Glu Glu Ala Val Val Lys Thr Val Val Gly Glu

420 425 430

Leu Asp Gln Leu Gly Ser Val Leu Ser Leu Arg Val His Ser Val Glu

435 440 445

Glu Lys Ser Ser Lys Ile Ser Asn Ile Thr Met Glu Gln Arg Leu Pro

450 455 460

Ser Lys Val Pro

465

<210> 49

<211> 7

<212> PRT

<213> Artificial sequence

<220>

<223> His6 tag

<400> 49

Met His His His His His His

1 5

Claims

1. A method for producing a multifilament, comprising a step of ejecting a spinning solution containing modified fibroin and a solvent from a spinning nozzle having a number of holes of 1000 or more and a pore diameter of 0.01mm to 0.6mm so as to contact the modified fibroin with a coagulation liquid and coagulating the modified fibroin,

The modified fibroin is of the formula 1: [ (a) _n motif-REP ] _m or formula 2: an altered fibroin having a domain sequence represented by the (A) _n motif-REP ] _m-(A)_n motif, wherein REP represents an amino acid sequence consisting of 2 to 200 amino acid residues, the (A) _n motif is an amino acid sequence having an alanine residue number of 2 to 27, the ratio of the alanine residue number in the (A) _n motif to the total number of amino acid residues is 40% or more, m represents an integer of 2 to 300,

The engineered fibroin comprises:

An engineered fibroin having a domain sequence with reduced glycine residue content;

An engineered fibroin having a domain sequence with reduced (a) _n motif content;

An engineered fibroin having a domain sequence with reduced glycine residue content and (a) _n motif content; or alternatively

An engineered fibroin having a domain sequence with reduced glutamine residue content,

In the case of an engineered fibroin having a domain sequence with reduced glycine residue content, the ratio of the motif sequence in which glycine residues are replaced with other amino acid residues is 10% or more with respect to the entire motif sequence,

In the case of an engineered fibroin having a domain sequence with reduced content of (A) _n motif, 10 to 40% of the amino acid sequence of (A) _n motif is deleted from the naturally-derived fibroin,

In the case of an engineered fibroin having a domain sequence with a reduced glutamine residue content, the glutamine residue content is 9% or less.

2. A multifilament produced by the method for producing a multifilament according to claim 1, which has 1000 or more constituent filaments, a coefficient of variation of elastic modulus of 15% or less, a coefficient of variation of strength of 15% or less, and a coefficient of variation of elongation of less than 33%.

3. A multifilament according to claim 2, wherein the elongation of the multifilament has a coefficient of variation of 20% or less.

4. A multifilament according to claim 2 or 3, wherein the elongation of the multifilament has a coefficient of variation of 10% or less.

5. A multifilament according to claim 2 or 3, wherein the coefficient of variation of the titer of the multifilament is 20% or less.

6. A multifilament according to claim 2 or 3, wherein the coefficient of variation in the strength of the multifilament is 10% or less and the coefficient of variation in the elastic modulus of the multifilament is 10% or less.

7. A multifilament yarn according to claim 2 or 3, wherein the modified fibroin has an average hydrophobicity index of more than-0.8.

8. A multifilament yarn according to claim 2 or 3, wherein the modified silk fibroin is a modified spider silk fibroin.

9. A multifilament yarn according to claim 2 or 3, wherein it has a shrinkage history after spinning which is irreversibly shrunk.

10. A multifilament yarn according to claim 9, wherein the shrinkage history is a shrinkage history by contacting the multifilament yarn with water to cause irreversible shrinkage or a shrinkage history by heat-relaxing the multifilament yarn to cause irreversible shrinkage.

11. A multifilament yarn according to claim 10, wherein the shrinkage of the multifilament yarn having a shrinkage history after spinning which is irreversibly shrunk is 5% or less, as defined by the following formula;

shrinkage [% ] = (1- (length of multifilament when dried from wet state/length of multifilament when wet state)) ×100.

12. A multifilament yarn according to claim 2 or 3, wherein the coefficient of variation of the titer of the multifilament yarn is 0.01% to 6.5%.

13. A multifilament yarn according to claim 2 or 3, wherein the multifilament yarn has a coefficient of variation in strength of 0.01% to 3.8%.